Characterization of a Small Family (CAIII) of Microsatellite-Containing Sequences with X-Y Homology

Four X-linked loci showing homology with a previously described Y-linked polymorphic locus (DYS413) were identified and characterized. By fluorescent in situ hybridization (FISH), somatic cell hybrids, and YAC screening, the X-linked members of this small family of sequences (CAIII) all map in Xp22, while the Y members map in Yq11. These loci contribute to the overall similarity of the two genomic regions. All of the CAIII loci contain an internal microsatellite of the (CA)_n type. The microsatellites display extensive length polymorphism in two of the X-linked members as well as in the Y members. In addition, common sequence variants are found in the portions flanking the microsatellites in two of the X-linked members. Our results indicate that, during the evolution of this family, length variation on the Y chromosome was accumulated at a rate not slower than that on the X chromosome. Finally, these sequences represent a model system with which to analyze human populations for similar X- and Y-linked polymorphisms. Received: 29 July 1996 / Accepted: 15 January 1997     

Transfection of Reaggregating Embryonic Chicken Retinal Cells with an Antisense 5'-DNA Butyrylcholinesterase Expression Vector Inhibits Proliferation and Alters Morphogenesis

Abstract: The function of the enzyme butyrylcholinesterase (BChE) in the developing and mature brain is still unclear. We have inserted 577 bp of the 5' upstream region plus 106 bp of the exon 1 of the rabbit BChE gene in reverse orientation under control of an SV40 early promoter derivative in an expression vector. This vector was introduced by calcium phosphate-mediated transfection into embryonic chicken retina cells during the first days of reaggregation culture. Depending on the retinal origin, the transfected cell population forms histotypic retina-like spheres, so-called rosetted or stratified retinospheroids. We show that antisense 5'-BChE gene expression decreased the steady-state mRNA level of BChE and the translation of the BChE protein, inhibited proliferation, and accelerated histogenesis in both cellular systems. The pronounced effects of antisense 5'-BChE transfection of spheroids document a key role of BChE during the early reaggregation process of retinal cells, most likely by regulating their growth and differentiation.     

Establishment and characterization of a new human hepatocellular carcinoma cell line

Summary A human hepatocellular carcinoma cell line (FOCUS—Friendship of China and United States) was derived from a patient with primary hepatocellular carcinoma. This cell line has been in continuous culture over an 18-mo period. The morphological and ultrastructural features of FOCUS are consistent with its neoplastic hepatocellular orgin. FOCUS cells contain aspartate aminotransferase and glucose-6-phosphatase activity. In addition, α₁-antitrypsin, fibrinogen, alpha fetoprotein, and carcinoembryonic antigens were detectble in the cytoplasm of the cultured cells by immunochemical staining techniques. The karyotype of the FOCUS cell is human in origin and it contains human DNA sequences as detected by molecular hybridization analysis. The FOCUS cells do not show evidence of density-dependent inhibition of growth under confluent conditions. Repeated growth curves over an 18-mo period were identical, revealing a doubling time of 42 to 48 h. The malignant potential of FOCUS cells was further demonstrated by their ability to lead to gross tumor formation after subcutaneous infection into nude mice. From one of the solid tumors grown in nude mice, recultured cell lines have been established and found to have properties identical to the original FOCUS cell line. This FOCUS cell line represents an additional model for further investigation of tumor specific antigens and the relationship between hepatitis B virus (HBV) and hepatocellular carcinoma. Preliminary molecular characterization has indicated the existence of integrated HBV sequences within the FOCUS genome.     

Erythrocytic cation transport receptor numbers and activity in pregnancies complicated by essential hypertension and pre-eclampsia.

Various functions of erythrocytic cation transport were studied in normotensive and hypertensive pregnancy (women with pre-eclampsia and essential hypertension). The results showed that in pregnancy there is an increase in the number of erythrocytic glycoside binding sites accompanied by a proportional increase in the active inward transport of rubidium (used as a substitute for potassium). There was no evidence of an effect of pregnancy on intraerythrocytic sodium concentrations. These changes were apparently entirely attributable to pregnancy and not affected by pre-eclampsia or essential hypertension. It is suggested that these alterations indicate an adaptive increase in sodium pump numbers and activity secondary to a tendency for the intraerythrocytic sodium concentration to rise during pregnancy and compensating for that tendency.     

Met-enkephalin immunoreactivity in blood platelets

Picogram amounts (50–150 pg/mg protein) of immunoreactive met-enkephalin material (met-enkephalin in IR) were detected by radioimmunoassay in human, rat and rabbit platelets. Characterization of this material by thin-layer chromatography, gel filtration chromatography and high-pressure liquid chromatography indicated that it behaves identically with synthetic met-enkephalin. No high molecular weight met-enkephalin IR could be detected in the platelet extracts, even after trypsin hydrolysis, using two antisera which are able to recognize some of the putative met-enkephalin precursors present in the adrenal gland or striatum. $\text{In vitro}$, thrombin released platelet met-enkephalin in IR concomitantly with 5-hydroxytryptamine (5-HT), suggesting a common subcellular localization, i.e. the 5-HT storing organelles, for met-enkephalin IR and the amine. $\text{In vivo}$, platelet met-enkephalin IR in the Sprague-Dawley rat was affected neither by adrenalectomy nor by hypophysectomy. Thirteen- and 18-week-old spontaneous hypertensive rats (SHR) had lower platelet concentrations of met-enkephalin in IR than age matched normotensive Wistar-Kyoto rats.     

Decreased sensitivity of human platelets to PGI2 during long-term intraarterial prostacyclin infusion in patients with peripheral vascular disease — A rebound phenomenon?

During successful treatment of peripheral vascular disease with synthetic prostacyclin no alteration in platelet function was reported (1).In 8 patients infused with synthetic prostacyclin continuously for 7 days intraarterially, the platelet function was monitored. Special attention was drawn to the platelet sensitivity in vitro for PGI₂, which is discussed as an important factor maintaining the hemostatic balance.In all the patients with peripheral vascular disease between 24 and 48 hours after the beginning of the infusion a sudden decrease in platelet sensitivity accompanied by an increase in platelet count could be seen.These dramatic alterations representing probably a rebound phenomenon occurring during long-term PGI₂-treatment might be an explanation for a non-beneficial effect of the treatment and in some cases a limiting factor for the continuation of the infusion itself. It is not clear, if this rebound phenomenon is due to a stimulation of an endogenous inhibitor, lowering the synthesis of a naturally occurring substance acting against this inhibitor or tachyphylaxia.     

Enhanced erythroid burst formation in mice after testosterone treatment

A single administration of testosterone propionate (TP) in ex-hypoxic polycythemic mice induces, at 24 hr after androgen, an amplification of the erythroid burst-forming unit (BFU-E or B) pool in marrow. This phenomenon is not associated with an amplification of the erythroid colony-forming unit (CFU-E or E) compartment and is followed by its depletion. In the other hand, the 36–49 hr rise of erythropoietin (Ep) levels in serum is followed by a 60-hr amplification of the E pool. It is suggested that the latter phenomenon is mediated by enhanced Ep production, whereas the early amplification of the B compartment may derive from a direct influence of TP at the stem cell level.     

Neoflavonoids and the cinnamylphenol kuhlmannistyrene from Machaerium kuhlmannii and M. Nictitans

The trunkwood of Machaerium kuhlmannii contains methyl palmitate, 3-O-acetyloleanolic acid and sitosterol; the benzene derivatives 2,3-dimethoxyphenol, 2,6-dimethoxyphenol, 2-hydroxy-3-methoxyphenol, 2,3-dimethoxybenzaldehyde and methyl 3-(2-hydroxy-4-methoxyphenyl)-propionate; the isoflavonoids formononetin and (6aS,11aS)-medicarpin; the neoflavonoids (R)-3,4-dimethoxydalbergione, (R)-3,4-dimethoxydalbergiquinol, kuhlmanniquinol [(R)-3-(4-hydroxyphenyl)-3-(5-hydroxy-2,3,4-trimethoxyphenyl)-propene], dalbergin, kuhlmannin (6-hydroxy-7,8-dimethoxy-4-phenylcoumarin) and kuhlmannene (6-hydroxy-7,8-dimethoxy-4-phenylchrom-3-ene), as well as the cinnamylphenol kuhlmannistyrene [Z-1-(5-hydroxy-2,3,4-trimethoxybenzyl)-2-(2-hydroxyphenyl)-ethylene]. Five of these compounds, in addition to (R)-4′-hydroxy-3,4-dimethoxydalbergione, were also isolated from a trunkwood extract of M. nictitans. Structural assignments were confirmed by chemical interconversion and by the synthesis of (±)-kuhlmanniquinol.     

Absolute configurations of isoflavans

The absolute configurations of isoflavans and isoflavanquinones isolated from Cyclolobium, Dalbergia and Machaerium species were established by comparison of their ORD curves with that of (3S)-5,7,3′,4′-tetra-methoxyisoflavan and (3S)-7,4′-dimethoxyisoflavan-2′,5′-quinone, respectively. The assignments were checked by the ozonolysis of the isoflavan (?)-duartin to (R)-paraconic acid and the oxidation of isoflavans to isoflavanquinones. The PMR spectra of the dihydropyran ring of the isoflavans are discussed in terms of the preferred conformation of this ring.