Effects of long-term fixation on histological quality of undecalcified murine bones embedded in methylmethacrylate

While long-term fixation and storage of specimens is common and useful for many research projects, it is particularly important for space flight investigations where samples may not be returned to Earth for several months (International Space Station) or years (manned mission to Mars). We examined two critical challenges of space flight experimentation: the effect of long-term fixation on the quality of mouse bone preservation and the preservation of antigens and enzymes for both histochemical and immunohistochemical analyses, and how the animal/sample processing affects the preservation. We show that long-term fixation minimally affects standard histological staining, but that enzyme histochemistry and immunolabeling are greatly compromised. Further, we demonstrate that whole animal preservation is not as suitable as whole leg or stripped leg preservation for long-term fixation and all histological analyses. Overall, we recommend whole leg processing for long-term storage of bone specimens in fixative prior to embedding in plastic for histological examination.     

柠檬酸合酶的分子生物学研究进展

柠檬酸合酶（citrate synthase,CS）是细胞内多种重要代谢途径的关键酶。CS可催化草酰乙酸和乙酰辅酶A之间的缩合反应生成柠檬酸和辅酶A。通常革兰氏阳性细菌、古菌以及真核细胞的CS为同源二聚体,而革兰氏阴性细菌的CS为同源六聚体。根据其在细胞内的定位不同,CS可分为线粒体CS、乙醛酸循环体CS、过氧化物酶体CS。这些同工酶在能量代谢、植物脂肪的代谢、脂肪酸的氧化及细胞解毒过程中起着重要作用。不同来源的CS空间结构、催化机制和动力学性质十分相似。针对其生化特性、空间结构特点、催化机制以及分子进化等研究进展进行综述。     

Variation in heat shock proteins within tropical and desert species of poeciliid fishes

The 70-kilodalton heat shock protein (hsp70) family of molecular chaperones, which contains both stress-inducible and normally abundant constitutive members, is highly conserved across distantly related taxa. Analysis of this protein family in individuals from an outbred population of tropical topminnows, Poeciliopsis gracilis, showed that while constitutive hsp70 family members showed no variation in protein isoforms, inducibly synthesized hsp70 was polymorphic. Several species of Poeciliopsis adapted to desert environments exhibited lower levels of inducible hsp70 polymorphism than the tropical species, but constitutive forms were identical to those in P. gracilis, as they were in the confamilial species Gambusia affinis. These differences suggest that inducible and constitutive members of this family are under different evolutionary constraints and may indicate differences in their function within the cell. Also, northern desert species of Poeciliopsis synthesize a subset of the inducible hsp70 isoforms seen in tropical species. This distribution supports the theory that ancestral tropical fish migrated northward and colonized desert streams; the subsequent decrease in variation of inducible hsp70 may have been due to genetic drift or a consequence of adaptation to the desert environment. Higher levels of variability were found when the 30- kilodalton heat shock protein (hsp30) family was analyzed within different strains of two desert species of Poeciliopsis and also in wild-caught individuals of Gambusia affinis. In both cases the distribution of hsp30 isoform diversity was similar to that seen previously with allozyme polymorphisms.      

Antitumour activities of levans produced by Zymomonas mobilis strains

Levans produced by four Zymomonas mobilis strains showed antitumour activity against sarcoma 180 and Ehrlich carcinoma in Swiss albino mice. Levans from two strains (ZAP and CP4) had the highest effects. NMR analysis showed that the polymers were composed only of fructose units. The results suggested that the antineoplasic effect is associated to the polysaccharide molecular weight and that a particular molecular weight range may be responsible for this effect.     

Impact of anaerobic digestion on organic matter quality in pig slurry

Changes in pig slurry organic matter (OM) during anaerobic digestion (AD) were studied in a reactor to characterize OM evolution through AD. OM maturity and stability were evaluated using different biological and physico-chemical methods. Germination and growth chamber experiments revealed a higher maturity of digested slurry (DS) than raw slurry (RS). Soil incubations showed that DS was more stable than RS with a C-mineralization of 12.0 g CO₂-C 100 g^?1 C_org after 49 days as compared to 17.6 g CO₂-C 100 g^?1 C_org. Biochemical fractionation showed a relative increase in stable compounds such as hemicellulose-like and lignin-like molecules. Fourier-transform infrared spectroscopy showed some changes in the chemical structures of OM with a reduction in the aliphatic chain, lipid and polysaccharide levels. A comparison between the evolution of OM during AD and the first weeks of a composting process showed almost identical changes. Finally a theoretical method called Fictitious Atomic-group Separation was applied to the elemental compositions of RS and DS. DS was less humified than RS and presented the properties of a fulvic acid, indicating that the observed stability in DS was mainly due to the biodegradation of the most labile compounds.     

Mitochondrial phylogeny of notothenioids: a molecular approach to Antarctic fish evolution and biogeography

Antarctic waters represent a unique marine environment delimited by an oceanographic barrier, the Polar Front Zone, and characterized by constant subzero temperatures and presence of sea ice. A group of teleost fish, the Notothenioidei, have adapted to these challenging environmental conditions, undergoing a remarkable diversification. In the present study a total of 798 base pairs, generated from partial sequencing of 16S and 12S mitochondrial ribosomal RNA genes, were examined in 33 notothenioid species representative of all families included in the suborder Notothenioidei. Phylogenetic trees, reconstructed on the basis of sequence data by different methods, indicate that traditional hypotheses on notothenioid systematics and biogeography might be in need of reexamination. Molecular evidence suggests that vicariant speciation could be invoked to explain the early divergence of Eleginops maclovinus, a species previously included in the family Nototheniidae, which is now proposed as the closest sister group to all the rest of notothenioids apart from bovichtids. On the other hand, repeated, independent dispersal through the Polar Front is proposed for the divergence of other subantarctic notothenioid species. Likewise, multiple, independent transitions from benthic to pelagic habit are inferred from molecular data, at variance with the more conservative hypothesis based on cladograms reconstructed from morphological data.     

Expression from cell type-specific enhancer-modified retroviral vectors after transduction: influence of marker gene stability

The enhanced green fluorescent protein (EGFP) is increasingly used as a reporter gene in viral vectors for a number of applications. To establish a system to study the activity of cis-acting cellular regulatory sequences, we deleted the viral enhancer in EGFP-carrying retroviral vectors and replaced it with cell type-specific elements. In this study, we use this system to demonstrate the activity of the human CD2 lymphoid-specific and the Tie2 endothelial cell type-specific enhancers in cell lines and in primary cells transduced by retroviral vectors. Furthermore, we compare findings obtained with EGFP as the reporter gene to those obtained replacing EGFP with d2EGFP, an unstable variant of EGFP characterized by a much shorter half-life compared to EGFP, and by reduced accumulation in the cells. d2EGFP-carrying vectors were generated at titers which were not different from those generated by the corresponding vectors carrying EGFP. Moreover, the activity of a Moloney murine leukemia virus enhancer could be readily detected following transduction of target cells with either EGFP- or d2EGFP-carrying vectors. However, the activity of the relatively weak CD2 and Tie2 enhancers was exclusively detected using EGFP as the reporter gene.These findings indicate that enhancer replacement is a feasible and promising approach to address the function of cell type-specific regulatory elements in retroviral vectors carrying the EGFP gene.