A nonradioisotope biomedical assay for intact oligonucleotide and its chain-shortened metabolites used for determination of exposure and elimination half-life of antisense drugs in tissue.

Rigorous extraction methods coupled with capillary gel electrophoresis (CGE) provide a basis for a nonradiolabel assay for quantitation of intact antisense drug and its numerous chain-shortened metabolites. As part of the validation of the CGE method, we compared the quantitation of unlabeled ISIS 3521 (ISI 641A) and its chain-shortened metabolites with total radioactivity of [(35)S]-ISIS 3521. ISIS 3521 was labeled on the fifth nucleotide linkage from the 5'-end with (35)S by well-established methods. Multiple tissues collected from rats after administration of [(35)S]-ISIS 3521 were assayed by both radiolabel (liquid scintillation spectroscopy) and CGE methods. The CGE method provided accurate quantitation of the drug and its metabolites in kidney cortex and liver tissues. The correlation between methods for multiple tissues over time was excellent with 88.5% of the measurements being statistically equivalent. These data suggest that CGE is an accurate means of quantitating oligonucleotide in tissue and that it compares favorably with traditional radiochemical techniques. Clearance half-lives for total measurable oligonucleotides were equivalent to clearance of total radioactivity in both liver and kidney with the longest clearance half-life associated with the kidney.     

Influence of development rate on pharyngeal teeth formula in <Emphasis Type="Italic">Abramis brama</Emphasis> (L.) bream: Experimental cata

An experiment on acceleration and retardation of ontogeny with thyroid manipulation has revealed direct changes in definitive dentition of pharyngeal bones in Abramis brama bream. As development rate accelerates, the number of teeth reduces to the formula 5-4. When development rate is retarded, this number increases to the formula 6-5. Moreover, an additional minor row of teeth (1.6–5.1, 2.6–5.2) is formed. The observed changes in tooth numbers exceed the known variability in natural populations of bream. It is assumed that heterochronies lead to the changes in the number of teeth.     

Significance of mirex-caused hypoglycemia and hyperlipidemia in rats

Treatment of rats with mirex (40 ppm in diet) caused hypoglycemia, liver enlargement, and inhibition of adrenal corticosteroid-synthesizing enzyme activity. At toxic dosages (20,000 ppm mirex in diet, which has a lethal toxicity-50 [LT-50] of ten days) poisoned female rats showed severe hypoglycemia, fatty liver, adrenal hyperplasia, hypophagia, lipid mobilization, and body weight (bw) loss. A 50 μg/kg intraperitoneal (IP) dose of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) in male rats caused similar effects two days posttreatment. Hypoglycemia could be overcome by prednisone (which also inhibited adrenocorticoid-synthesizing enzyme activities) but not by streptozotocin treatment, indicating that hypoglycemia may be related to glucocorticoid deficiency resulting from inhibition of their synthesis and not by direct effects on pancreatic β-cells. Glucocorticoid deficiency could also cause increased release of adrenocorticoid hormone (ACTH), which may enhance fat mobilization caused by hypophagia.     

Dimorphism in methane seep-dwelling ecotypes of the largest known bacteria

We present evidence for a dimorphic life cycle in the vacuolate sulfide-oxidizing bacteria that appears to involve the attachment of a spherical Thiomargarita-like cell to the exteriors of invertebrate integuments and other benthic substrates at methane seeps. The attached cell elongates to produce a stalk-like form before budding off spherical daughter cells resembling free-living Thiomargarita that are abundant in surrounding sulfidic seep sediments. The relationship between the attached parent cell and free-living daughter cell is reminiscent of the dimorphic life modes of the prosthecate Alphaproteobacteria, but on a grand scale, with individual elongate cells reaching nearly a millimeter in length. Abundant growth of attached Thiomargarita-like bacteria on the integuments of gastropods and other seep fauna provides not only a novel ecological niche for these giant bacteria, but also for animals that may benefit from epibiont colonization.     

酸敏感离子通道与脑梗死

急性脑梗死约占全部脑卒中的70%,病死率和致残率高,且极易复发。但目前针对急性脑梗死在时间窗内溶栓、抗凝等治疗手段不能从根本上切实有效地修复受损脑组织,且伴有出血等风险。寻找脑梗死形成发展的原因并予以治疗迫在眉睫。酸中毒是引起缺血性脑损伤的重要机制。大量实验研究表明,酸中毒能加重神经元的缺血性损伤,且其梗死面积与酸中毒的程度直接相关。但缺血产生的酸中毒如何引起神经元损伤的确切机制尚不明确。最近研究发现酸中毒能激活一种在中枢及周围神经中广泛存在的膜通道,即酸敏感离子通道,它对Ca2+通透,能引起细胞内Ca2+超载,同时能激活胞内酶引起细胞内蛋白质、脂类及核酸的降解,加重缺血后脑损伤。本文就酸敏感离子通道1a与脑梗死做一综述。     

Comparison of the tissue-specific expression and developmental regulation of two closely linked rodent genes encoding cytosolic retinol-binding proteins

Cellular retinol-binding protein (CRBP) and cellular retinol-binding protein II (CRBP II) are two highly homologous cytoplasmic proteins that bind all-trans-retinol. We have recently demonstrated that the mouse genes encoding CRBP and CRBP II are closely linked on chromosome 9 and that both human genes are located on chromosome 3 (Demmer, L.A., Birkenmeier, E.H., Sweetser, D.A., Levin, M.S., Zollman, S., Sparkes, R.S., Mohandas, T., Lusis, A.J., and Gordon, J.I. (1987) J. Biol. Chem. 262, 2458-2467). We have now used RNA blot hybridization analysis to assess the degree to which these genes are coordinately expressed in fetal, suckling, weaning, and adult rat tissues. Both genes exhibit different developmental patterns of expression in liver, intestine, lung, kidney, testes, and placenta. In the intestine, CRBP mRNA was detected during the 16th day of gestation--prior to the development of a well-differentiated absorptive epithelium--and remained essentially unchanged throughout the peri- and postpartum periods. By contrast, the pattern of intestinal CRBP II mRNA accumulation closely parallels the times of first appearance, and subsequent proliferation, of the intestinal absorptive columnar epithelium, supporting the hypothesis that CRBP II is involved in the intestinal uptake or intracellular trafficking of this hydrophobic vitamin. In the fetal liver, both genes were expressed by gestational day 16. Whereas the concentration of hepatic CRBP mRNA increased markedly during the suckling and early weaning periods, CRBP II mRNA levels fell abruptly immediately after birth. These peripartum changes were not paralleled by remarkable alterations in the steady state levels of hepatic retinol. Marked changes in the expression of CRBP in the liver and of CRBP II in the intestine were also documented in pregnant and lactating female rats. These differences in CRBP/CRBP II gene expression strongly suggest that their proteins serve different physiological functions. The peripartum liver may provide a useful model for dissecting the relative roles played by these homologous proteins in retinoid metabolism as well as the factors which modulate activation and repression their genes.     

Photoaffinity labeling of T4 bacteriophage 32 protein

With a view toward the determination of nucleic acid binding domains and sites on nucleic acid helix-destabilizing (single strand-specific) proteins (HDPs), we have studied the interactions of the copolymer polynucleotide photoaffinity label, poly(adenylic, 8-azidoadenylic acid), (poly(A,8-N3A] with the T4 bacteriophage HDP, 32 protein. Poly(A,8-N3A) quenched the intrinsic tryptophan fluorescence of 32 protein in a manner similar to that observed with other polynucleotides, and the effect could be reversed by addition of sufficient NaCl. The binding affinity and site size of this noncovalent interaction of poly(A,8-N3A) with 32 protein are similar to the values obtained for poly(A) and this protein. When [3H]poly(A,8-N3A)/32 protein mixtures were irradiated at 254 nm, fluorescence quenching was not reversed by NaCl, suggesting that the label was covalently bound to the protein. Mixtures of photolabel and protein subjected to short periods of irradiation (generally 1 min, 2000 erg mm-2) formed high molecular weight complexes, which when electrophoresed on sodium dodecyl sulfate (SDS)-polyacrylamide gels were radioactive and stained with Coomassie Blue R. Under the same conditions, [3H]poly(A) failed to label 32 protein. The radioactivity of [3H]poly(A,8-N3A)-labeled complexes subjected to micrococcal nuclease after irradiation was seen to migrate just behind the free 32 protein monomer on SDS-polyacrylamide gels, indicating that portions of the photolabel not in direct contact with protein were accessible to this enzyme. By several criteria, we conclude that 32 protein was photolabeled specifically at its single-stranded nucleic acid binding site. Single-stranded nucleic acids with affinities for protein greater than that of poly(A,8-N3A) effectively inhibited photolabeling. The [NaCl] dependence of photolabeling monitored on SDS gels paralleled the NaCl reversal of (noncovalent) poly(A,8-N3A)-32 protein binding. Photolabeling reached a plateau after 1-2 min. The formation of high molecular weight complexes with increasing [poly(A,8-N3A)] paralleled the disappearance of free protein on SDS gels, and reached a saturation level of about 75% labeling. Several chromatographic procedures appear to be useful for the separation of the photolabeled complexes from free protein and photolabel. Limited trypsin hydrolysis of photolabeled 32 protein indicated that all the label was within the central ("III") portion of the protein. This approach should have general applicability to the identification of nucleic acid binding sites on helix-destabilizing proteins.     

黑龙江省三江平原丹顶鹤的数量分布

在地面调查的基础上,我们使用Y-11轻型飞机对黑龙江省三江平原地区的丹顶鹤的数量分布近行了调查,调查时飞行高度80米,航速140公里/小时,续航里程共3748公里。调查结果表明,丹顶鹤在三江平原主要分布在8个地区,其中嘟噜河下游、洪河自然保护区、七星河流域和兴凯湖低地是主要繁殖地,总数量共309只。