RNA polymerase II transcription complex assembly in nuclear extracts

Variation in superovulatory responses in cattle may be related to the stage of follicular growth at the time of gonadotropin treatment. Waves of follicle growth are regulated by both follicle-stimulating hormone (FSH) and oestradiol. The objective of experiment 1 was to determine the dynamics of follicle wave emergence and the relationship with FSH and oestradiol concentrations, after treatment of heifers with oestradiol benzoate (ODB) in the presence of an intravaginal progesterone-releasing device (CIDR-B). Experiment 2 examined the superovulatory response, embryo yield and quality following treatment with porcine follicle-stimulating hormone (pFSH) at different times relative to ODB injection. In experiment 1, 28 beef heifers were treated with a CIDR for 9 days and allocated at random to one of four groups to receive either: (I) CIDR only, or 5 mg ODB given as a single intramuscular injection at (II) day 0 (d0); (III) day 1.5 (d1.5); or (IV) day 3 (d3) post CIDR insertion. Ovaries were examined using daily ultrasound and blood samples were collected twice daily for 11 days. In experiment 2, 96 heifers were treated with a CIDR and 5 mg ODB as in experiment 1, and were allocated using a 4 x 3 factorial design plan to a superovulation programme using three doses (400 IU; 600 IU; 800 IU) of pFSH. FSH was given for 4 days at 12-h intervals beginning 6.5 days after CIDR insertion. Heifers received prostaglandin analogue 12 h before CIDR removal and were inseminated (AI) at 48 and 60 h post CIDR withdrawal and embryos were recovered 7 days after AI. In experiment 1, the interval from CIDR insertion to follicle wave emergence (FWE) was longer (P < 0.05) in heifers treated with ODB at d1.5 (5.4 +/- 0.4 days) and d3 (5.1 +/- 0.6 days) compared to heifers treated with CIDR only (2.4 +/- 0.4 days). On the basis of time to proposed injection of pFSH heifers would have had follicle emergence 4.4, 2.3, 1.5 and 1.4 days prior to pFSH for groups I, II, III and IV, respectively. In experiment 2, heifers treated with ODB at d1.5 had a higher (P < 0.05) superovulatory response (18.2 +/- 1.7) than heifers treated at d3 (12.8 +/- 1.7), but superovulatory response in both groups did not differ (P > 0.05) from heifers treated at d0 (14.4 +/- 2.0) or with CIDR only (15.0 +/- 1.8). There were fewer (P < 0.05) freezable-grade embryos recovered from heifers treated with ODB at d0 (1.5 +/- 0.7) and d3 (2.1 +/- 0.5) compared to heifers treated at d1.5 (3.0 +/- 0.6) or in heifers treated with CIDR only (3.4 +/- 0.7). Increasing the dose of pFSH caused a linear increase in the superovulatory response (11.7 +/- 1.0, 15.8 +/- 1.4 and 18.0 +/- 1.9) and in the number of embryos recovered (5.8 +/- 0.9, 7.0 +/- 0.8 and 9.1 +/- 1.0) for 400 IU, 600 IU and 800 IU, respectively. In conclusion, heifers treated with ODB had wide variation in time to follicle wave emergence and there was not a consistent beneficial effect of pretreatment with ODB on embryo yield and quality following superovulation.     

RNA polymerase II transcription initiation: a structural view

The emergence of new strains of Vibrio Cholerae has added a new dimension to the variability in pathogenicity and potential virulence of the organisms precipitating diarrhoeal diseases. Considering the shifting patterns of V. cholerae 01 there is a continuous need to monitor the strain characteristics. In this study total 541 stool specimens of acute secretory diarrhoea were investigated between May 1992 and November 1994 for strains of Vibrio Cholerae and anti-microbial susceptibility testing of all the confirmed V. Cholerae strains. In 1992, 50 of the 125 strains (40%) were positive for V. cholerae 01 predominantly biotype El Tor serotype ogawa, and 10 (80%) of non 01 type, with most strains susceptible to tetracycline (100%), chloramphenicol (98%) and Cotrimoxazole (98%), but all resistant to polymyxin B and furazolidine. In 1993, 44 (43.6%) of the 010 strains were positive for V. cholerae 0139 and the rest V. cholerae 01. In 1994, another sero group of V. cholerae 010 emerged, with 42 (13.3%) being positive. Isolates did not agglutinate with any of these antisera and have been labelled as 'other than non-01 vibrio cholerae'.     

The role of activators in assembly of RNA polymerase II transcription complexes

Since the negative results of the international Bypass Study, extracranial-intracranial (EC-IC) bypass surgery is infrequently employed in the treatment of patients with cerebral ischemia. Newly acquired evidence concerning the pathophysiology of cerebral ischemia, however, has facilitated the identification of a small subgroup of patients with "hemodynamic" cerebral ischemia. Characteristically, these patients demonstrate severely impaired cerebrovascular reserve capacity due to occlusive disease and insufficient collateral blood supply. Over an 8-year period, 28 patients were defined by clinical and laboratory criteria as suffering from hemodynamic cerebral ischemia. All patients had recurring episodes of focal cerebral ischemia due to unilateral internal carotid artery occlusion. Computerized tomography (CT) scans either were normal or showed evidence of border zone infarction. The cerebrovascular reserve capacity was studied using 133Xe single-photon emission CT and acetazolamide challenge and was found to be significantly impaired in all patients. Based on these criteria, superficial temporal artery-middle cerebral artery anastomosis was performed to augment collateral flow to the ischemic hemispheres. Two patients died from myocardial infarction, one 4 days and the other 2 months postoperatively. One patient died from massive brain infarction and another suffered a postoperative stroke with incomplete recovery, resulting in a major morbidity and mortality rate of 14%. Minor morbidity included one patient with a subdural hematoma who subsequently recovered completely. The postoperative course was uneventful in 23 patients (82%). Over a mean follow-up period of almost 3 years, no patient had another episode of brain ischemia. Bypass patency was confirmed by postoperative angiography in 26 patients. Follow-up studies of cerebral blood flow (CBF) and cerebrovascular reserve capacity showed significant improvement of the latter while the resting CBF was essentially unchanged. In view of these findings, the authors conclude that EC-IC bypass surgery constitutes appropriate therapy for a subgroup of patients with recurrent focal cerebral ischemia, defined using the strict selection criteria employed in this study.     

Effect of hexachlorocyclohexane on hsp26 expression in transgenic Drosophila melanogaster

The type and amount of lipophilic antioxidants in plasma of hyperlipidemic patients are of great importance since they play a central role in preventing deleterious oxidation of blood lipids and proteins. Isolation and quantitation of lipophilic antioxidants from hyperlipidemic plasma samples meet great obstacles because of increased levels of various intermediary lipid products. This study was designed to develop a rapid and efficient extraction and separation procedure for simultaneous analysis of ubiquinone-9 and -10 as well as alpha-, delta-, and gamma-tocopherol isomers. The levels of ubiquinone-10, alpha- and gamma-tocopherol were analyzed in human plasma samples using high-performance liquid chromatography. Lipid extraction was performed by petroleum ether/methanol/water. After phase separation, ubiquinone was reduced to ubiquinol by sodium borohydride and the lipids were separated on a C18 column. A binary gradient with solvents containing lithium perchlorate was used, and an electrochemical detector was employed for quantitation. This procedure was also efficient for the analysis of antioxidant lipids in samples containing a large number of accumulated and interfering lipid intermediates. Thus, the procedure described here is useful for efficient and rapid quantitation of ubiquinones and tocopherols in human plasma samples, especially those originating from hyperlipidemic patients.     

Selection and analysis of rare second-site suppressors of Drosophila RNA polymerase II mutations

The disruptive effects on the activity-rest cycle of the monoamine oxidase inhibitor (MAOI) clorgyline and of continuous light were examined in Syrian hamsters. When administered in dim and moderate light intensities, clorgyline delayed the daily onset of wheel-running. When administered in bright light, it dissociated the circadian rhythm of wheel-running. This dissociation was prevented by lesions of the intergeniculate leaflet of the ventral lateral geniculate nucleus. Constant darkness restored the circadian rhythm of wheel-running in hamsters with disrupted circadian rhythms. The phase of the restored rhythm of wheel-running was shifted 6-12 h later than the phase of wheel-running prior to dissociation. Our results suggest that MAOI treatment weakens the coupling between oscillators that comprise the circadian pacemaker, and augments the disruptive effects of continuous light acting via the intergeniculate leaflet region of the ventral lateral geniculate nucleus. These effects on the circadian pacemaker may be responsible for disruptions of the sleep-wake cycle that occur as side effects when MAOIs are used clinically to treat depression and might play a role in the induction of mania and rapid cycling by antidepressants.     

Competent transcription initiation by RNA polymerase II in cell-free extracts from xeroderma pigmentosum groups B and D in an optimized RNA transcription assay

99Tcm-glucarate accumulation in human mammary BT-20 tumours hosted in severe combined immune deficiency (SCID) mice was compared to 111In-monoclonal antibody 103D2-F(ab')2, 99Tcm-methoxyisobutyl isonitrile (99Tcm-MIBI) and 99Tcm-diethylenetriamine pentaacetate (99Tcm-DTPA). The intracellular tumour distribution of 99Tcm-glucarate was also determined. SCID mice injected with a mixture of 99Tcm-glucarate and 111In-103D2-F(ab')2 were imaged serially up until 24 h. Computer planimetered tumour-to-blood activity (in the heart) ratios (T/BH) to 8 h were significantly greater for 99Tcm-glucarate than 111In-103D2. The mean (+/-S.D.) tumour-to-blood ratio (T/B) from biodistribution was 1.21 +/- 0.31 and 0.35 +/- 0.06 (P < 0.0001) at 5 h, and 1.526 +/- 0.29 and 0.75 +/- 0.2 (P < 0.0001) at 8 h, for 99Tcm-glucarate and 111In-103D2 respectively. At 24 h, T/B for 111In-103D2 (1.76 +/- 0.22) exceeded that of 99Tcm-glucarate (1.44 +/- 0.2, P = 0.01). 99Tcm-glucarate uptake in the tumours at 5 h (1.133 +/- 0.25 %ID g-1) and 8 h (1.213 +/- 0.23 %ID g-1) was significantly greater than that of 99Tcm-MIBI (0.340 +/- 0.09, P = 0.0002; 0.220 +/- 0.04, P = 0.0001) and 99Tcm-DTPA (0.091 +/- 0.03, P < 0.0002; 0.016 +/- 0.01, P < 0.0001) respectively. Intracellular tumour distribution of 99Tcm-glucarate was 50.91 +/- 6.55% in the nuclear fraction, 34.34 +/- 2.88% in the cytoplasmic fraction and 14.75 +/- 7.66% in the mitochondrial fraction. Thus glucarate may provide a 99Tcm-based mammary tumour imaging modality for visualization of tumours very quickly after tracer administration with maximal targeting in the nuclei of the tumour cells.