An in situ assessment of the routes and extents of colonisation of the mouse embryo by embryonic stem cells and their descendants

An embryonic stem (ES) cell line stably expressing lacZ under the control of an endogenous promoter has been isolated and used as a marker to follow the fate of ES cells injected into blastocysts and morulae, before midgestation. The results show a multisite pattern of blastocyst colonization by ES cells deposited into the blastocoel cavity and a low degree of mingling between ES cells and ICM cells. Furthermore, analysis of dispersal of ES cell descendants in postimplantation chimaeric embryos showed that colonization can be highly variable from one region of the embryo to another. In contrast, a high and reproducible degree of chimaerism was obtained when the ES cells were injected at the morula stage prior to ICM formation.     

Vessel Dimensions in Premature Atheromatous Disease of Aortic Bifurcation

The area ratio of the abdominal aortic bifurcation is defined as the ratio of the sum of the cross-sectional areas of the common iliac arteries to the cross-sectional area of the aorta. Abnormality of the area ratio is associated with initiation of atheroma of the lower aorta. Hypoplasia of the abdominal aorta predisposes to early occlusion as the atheromatous process advances.In five out of six women with thrombosis at the aortic bifurcation the area ratio was low. Hypoplasia of the abdominal aorta was present in two of these patients. All six patients were successfully treated by aortoiliac disobliteration and reconstruction.     

A structural basis for the biosynthesis of the major chlorogenic acids found in coffee

Chlorogenic acids (CGAs) are a group of phenolic secondary metabolites produced by certain plant species and an important component of coffee (Coffea spp.). The CGAs have been implicated in biotic and abiotic stress responses, while the related shikimate esters are key intermediates for lignin biosynthesis. Here, two hydroxycinnamoyl-coenzyme A shikimate/quinate hydroxycinnamoyl transferases (HCT/HQT) from coffee were biochemically characterized. We show, to our knowledge for the first time, that in vitro, HCT is capable of synthesizing the 3,5-O-dicaffeoylquinic acid diester, a major constituent of the immature coffee grain. In order to further understand the substrate specificity and catalytic mechanism of the HCT/HQT, we performed structural and mutagenesis studies of HCT. The three-dimensional structure of a native HCT and a proteolytically stable lysine mutant enabled the identification of important residues involved in substrate specificity and catalysis. Site-directed mutagenesis confirmed the role of residues leucine-400 and phenylalanine-402 in substrate specificity and of histidine-153 and the valine-31 to proline-37 loop in catalysis. In addition, the histidine-154-asparagine mutant was observed to produce 4-fold more dichlorogenic acids compared with the native protein. These data provide, to our knowledge, the first structural characterization of a HCT and, in conjunction with the biochemical and mutagenesis studies presented here, delineate the underlying molecular-level determinants for substrate specificity and catalysis. This work has potential applications in fine-tuning the levels of shikimate and quinate esters (CGAs including dichlorogenic acids) in different plant species in order to generate reduced or elevated levels of the desired target compounds.     

Interest of colchicine for the treatment of cystic fibrosis patients. Preliminary report

Cystic fibrosis (CF) lung disease is characterized by persistent inflammation. Antiinflammatory drugs, such as corticosteroids and ibuprofen, have proved to slow the decline of pulmonary function although their use is limited because of frequent adverse events. We hypothesized that colchicine could be an alternative treatment because of its antiinflammatory properties and upregulatory effect on cystic fibrosis transmembrane regulator (CFTR) closely related proteins. We herein present results obtained in an open study of eight CF children treated with colchicine for at least 6 months. Clinical status was better in all patients and respiratory function tests significantly improved in five. Median duration of antibiotherapy decreased significantly. These preliminary results support our hypothesis of a beneficial effect of colchicine in CF patients and stress the need for a controlled therapeutic trial.     

Conformational analysis of the adduct cis-[Pt(NH3)2 d(GpG)]+ in aqueous solution. A high field (500-300 MHz) nuclear magnetic resonance investigation.

A 500, 400 and 300 MHz proton NMR study of the reaction product of cis-Pt(NH3)2Cl2 or cis-[Pt(NH3)2 (H2O)2] (NO3)2 with the deoxydinucleotide d(GpG): cis-[Pt(NH3)2 d(GpG)] was carried out. Complete assignment of the proton resonances by decoupling experiments and computer simulation of the high field part of the spectrum yield proton-proton and proton-phosphorus coupling constants of high precision. Analysis of these coupling constants reveal a 100% N (C3'-endo) conformation for the deoxyribose ring at the 5'-terminal part of the chelated d(GpG) moiety. In contrast, the 3'-terminal -pG part of the molecule displays the normal behaviour for deoxyriboses: the sugar ring prefers to adopt an S (C2'-endo) conformation (about 70%). Extrapolating from this model compound, it is suggested that Pt chelation by a -dGpdG- sequence of DNA would require a S to N conformational change of one deoxyribose moiety as the main conformational alteration and lead to a kink in one strand of the double-helical structure of DNA.     

DNA fragment conformations IV - Helix-coil transition and conformation of d-CCATGG in aqueous solution by 1H-NMR spectroscopy.

The helix-coil transition and conformation of d-CCATGG were investigated using 1H-NMR spectroscopy at various frequencies (90, 276, 400 MHz). The changes in the chemical shifts and linewidths of imino protons between 5 degrees and 35 degrees C show that the d-CCATGG fraying process consists of two stages: the external dC.dG base pairs open at first, th internal dC.dG and central dA.dT base pairs then open simultaneously at higher temperatures similar to the case of d-ACATGT. The midpoint temperatures, the helix and coil proportions and the dissociation constant were determined from the sigma = f(t degree) curves of the base and sugar protons. The results indicate that the midpoint temperature increases with the number of the dG.dC base pair in a given size sequence, while the dissociation enthalpy appears to be independent. The difference between the T1 value of a base proton of the external and internal residues of the same nature is found to be a good criterion for base proton assignment. The high predominance of the S conformation for all residues shows that d-CCATGG duplexes adopt the B-helical conformation.     

Down-regulation of AP1 activities after polarization of vas deferens epithelial cells correlates with androgen-induced gene expression

Vas deferens epithelial cell subcultures were used to study the sequential regulation of jun/fos proto-oncogene expression and AP1 activities during cell proliferation, polarization and androgen-induced expression of a terminal differentiation marker, i. e. the mvdp gene. Proliferation of epithelial cells is associated with a high expression in the nucleus of most Jun and Fos oncoproteins. After cell seeding on an extracellular matrix which allows polarization and expression of the mvdp gene in response to androgens, AP1 protein accumulation is greatly altered and consists in a loss of JunB, Fra1, FosB and a decrease in c-Fos, c-Jun and Fra2, while JunD remained at the same level. This was correlated with a drop in AP1 binding activity as evaluated by gel shift assay using either AP1 consensus sequence or AP1 binding sites of the mvdp gene promoter region, and in AP1 transactivating activity, as estimated by stable transfection experiments using an AP1 responsive promoter (TRE-TK-luc). Androgens did not significantly influence AP1 activities. On the contrary, stimulation of AP1 proteins by the tumor-promoting phorbol ester caused a decrease in androgen-induced mvdp mRNA accumulation, and this effect was reversed by staurosporine, a potent inhibitor of PKC. Our data suggest that a down-regulation of AP1 activities induced by epithelial cell differentiation is a prerequisite to androgen-induced mvdp gene expression. The high AP1 activities observed during proliferative state or induced in TPA-treated polarized cells, exert a repressive effect on androgen action.     

三种裸趾虎核型及银带的研究

裸趾虎属Ｃｙｒｔｏｄａｃｔｙｌｕｓ动物 ,已知有 6 0余种 (Ｗｅｒ ｍｕｔｈ ,196 5 ) ,分布于亚州南部及西南部、欧州及大洋洲。中国记载有 5种 (Ｚｈａｏ等 ,1993) ,分布于西北部的干旱荒漠区及西藏东南部。有关其染色体研究报道甚少 ,仅国外记载 7种 (Ｓｚｃｚｅｒｂａｋ等 ,1996 ) ,且所提供的数据较为粗略。为此 ,本文对长裸趾虎Ｃ ｅｌｏｎｇａｔｕｓ、墨脱裸趾虎Ｃ ｍｅｄｏｇｅｎｓｉｓ和西藏裸趾虎Ｃ ｔｉｂｅｔａｎｕｓ核型进行研究 ,并首次报道其银带。旨在为分类及系统进化学提供细胞遗传学信息。1　材料和方法实验动物见表 1…