摘要:

Summary—The main limitation of non‐viral gene transfer methods is their relatively low efficiencyin vivo. However, a number of approaches can be taken to improve their performances, whether the aim is studying gene function during development or employing these techniques for gene therapy. Three non‐viral delivery systems that we have been particularly involved in in developing are described: the cationic lipid, dioctadecylamidoglycylspermine (DOGS), the cationic polymer polyethylenimine (PEI) and free DNA. The application of each of these methods to differentin vivosituations is presented: the use of DOGS for transfecting embryos and the developing mammalian nervous system; the recent application of PEI to the nervous system; and how naked DNA can be employed for transfecting different muscles and brain. The relative efficiencies are compared on the basis of luciferase reporter gene expression assessed in each tissue with the most appropriate vector system. Finally, the perspectives for constructing composite vectors combining safety and efficiency are considered br

ISSN:0248-4900    

DOI:10.1111/j.1768-322X.1995.tb00937.x

出版商:Blackwell Publishing Ltd    

年代:1995

数据来源: WILEY

摘要:

Summary—In contrast to general belief, the response of rabbit muscles to denervation is maturation to slow‐like type muscles [7]. We report now an investigation by biochemical, morphological, and mechanical studies of the time course effects of muscle denervation on the slow‐type soleus and fast‐type gastrocnemius to help clucidate the mechanism of maturation of rabbit denervated muscles to slow‐like muscles. In both muscles, denervation induced selective progressive atrophy of most fast fibers and hypertrophy of many slow fibers which displayed wide Z‐lines; this was accompanied by the appearance of hybrid LC1F‐ and LC1E‐associated slow myosins. The percentage of slow myosins increased with age similarly in the contralateral and denervated soleus. On the other hand, the percentage of slow myosins remained low in the contralateral gastrocnemius, whereas it increased to 95% in the denervated gastrocnemius; in the denervated gastrocnemius, the percentage of slow myosins reached 50% at about 35 days postnatal. At this age, the maximal shortening velocity of the denervated gastrocnemius and its twitch contraction time were already those of a slow‐type muscle. This suggests that in addition to myosin, other proteins contributed to the mechanical properties of the denervated gastrocnemius. Transformation of rabbit denervated muscles to slow‐like type muscles, which are associated with a lower energy requirement and higher muscle endurance than fast‐type muscles, may constitute an adequate model for human ne

ISSN:0248-4900    

DOI:10.1111/j.1768-322X.1995.tb00938.x

出版商:Blackwell Publishing Ltd    

年代:1995

数据来源: WILEY

摘要:

Summary—Immunohistochemical and physiological studies were carried out onNereis (Hediste) diversicolorOF Müller in order to obtain evidence concerning the neuroendocrine control of polychaete osmoregulation. The occurrence in this animal of peptides immunologically related to mammalian angiotensin II and I (AII and AI) and oxytocin (OT) was demonstrated in the brain and the ventral nerve cord (VNC) perikarya and nerve fibres as well as in a few peripheral structures (peripheral nerves, epithelial cells, nuchal organ, intestine and nephridia). The exact localization of immunoreactive cells was achieved by serial sections of brain and ventral nerve cord followed by a three‐dimensional reconstruction of brain ganglionic nuclei using the CATIA (‘Conception Assistée Tridimensionnelle Inter Active’) Dassault system program. Injections of polyclonal antisera against AII or OT provoked a partial inhibition of the increase in body weight inNereisexposed to hypo‐osmotic medium. The effect of a‐AII seemed more pronounced than that of a‐OT. In a subsequent test, injections of synthetic AII and AII‐amide (peptide recently isolated from an achaete (Salzetet al(1995)J Biol Chem270, 1575–1582) enhanced the increase in body weight and, therefore, strenghthened the hypothesis of the neuroendocrine control ofNereisosmoregulation. The antidiuretic effect of both synthetic peptides in this study was indicative of the exact role ofNereisendogenous melecule(s). AII was less potent than its amidated form. If AI‐like can easily be struck off the list of putative endogenous osmoregulaory factors, the role of OT‐like substance inNereisosmoregulation, which is partially demonstrated in this study, needs to be clarified by further physiological experiments using injection of synthetic peptide(s) or endogenous substance(s). All these results are discussed and compared to those recently obtained in an achaete annelid (Salzetet al(1993)Brain Res631, 247–255; Salzetet al(1993)Brain Res601, 173–184; Salzetet al(199

ISSN:0248-4900    

DOI:10.1111/j.1768-322X.1995.tb00939.x

出版商:Blackwell Publishing Ltd    

年代:1995

数据来源: WILEY

摘要:

Summary—The role of intestinal eosinophilic granule cells (EGCs) is still a subject of discussion. The aim of this study was to obtain additional functional data for a better characterization of these cells. Biochemical studies indicated the presence of small amounts of histamine, a characteristic and consistent marker of mast cells, in the posterior gut. On the other hand, histamine is always absent from homogenates of isolated EGCs. Using colorimetric assays, we were able to show aryl sulphatase B activity (18.5 ± 3.7 nM nitrocatechol/106cells) and detected peroxidase (1.86 ± 0.03 ng/106cells) in EGC homogenates. A cytochemical study enabled us to localize peroxidase in the granules of EGCs. These cells can also phagocytose latex beads. EGCs should thus be considered as homologous with mammalian eosinophils and not with mast cells. The screening for cells in the mucosae containing chondroitin sulphate revealed sparsely represented cells in the loose connective tissue in immediate proximity to blood capillaries. These cells could be mucosal mast ce

ISSN:0248-4900    

DOI:10.1111/j.1768-322X.1995.tb00940.x

出版商:Blackwell Publishing Ltd    

年代:1995

数据来源: WILEY

摘要:

Summary—K+accumulation‐depletion (AD) phenomena were found in single guinea pig ventricular myocytes using the patch‐clamp method in whole cell configuration. We suggest that the cardiomyocyte transverse‐axial tubular system (TATS) lumen is the restricted extracellular space where the K+AD could take place. A three‐dimensional (3D) reconstruction of the TATS in a cardiomyocyte segment from serial ultrafine sections was made by three‐dimensional isosurface rendering and quantitative data were obtained from the image processing. This original approach of the TATS intricated network gave a new vision of this membrane system; moreover, quantitative data about the tubular membrane importance (52.6% of the total plasma membrane) and its surface areaversusthe tubular volume fraction (STATS/VTATS= 13.5 μm2/μm3would fit in the electrophysiological results. The hypothesis whereby this ‘extracellular’ compartment could play, in single cells, a role as important as that of narrow clefts in the whole h

ISSN:0248-4900    

DOI:10.1111/j.1768-322X.1995.tb00941.x

出版商:Blackwell Publishing Ltd    

年代:1995

数据来源: WILEY

摘要:

Summary—We have previously demonstrated that intestinal and kidney finite cell lines were resistant toL monocytogenesinvasion (ieallowed low bacterial entry and no intracellular multiplication) in contrast to the continuous cell lines which were susceptible toListeriainvasion (ieallowed high bacterial entry and intracellular multiplication) (Velgeet al(1994a)Med Microbial Immunol183, 145). The aim of this study was to discover whether epigenetic or genetic cellular modifications could convertL monocytogenesresistant cells into a susceptible phenotype and to determine the cellular steps involved inListeriasusceptibility. Among the 5‐azacytidine treated finite cell lines, the untransformed immortal cell lines established remained resistant toL monocytogenesinvasion whereas the weakly transformed continuous cell lines established were converted into a susceptible phenotype. Transfection of resistant cells by SV40 large T antigen induced only highly transformed continuous cell lines displaying a susceptible phenotype. Taken together these data show that cell transformation enhancedListeriainvasion. This conclusion was supported by the observation thatL monocytogeneswas able to induce cell foci within murine finite cell monolayers. This morphological cell transformation was completely reversible and required live bacteria inside cells. In conclusion, we may speculate that theL monocytogenesintracellular multiplication observed within cell foci could be explained by the loss of contact inhibition of the finite cell monolayer. Indeed, the loss of both contact inhibition and anchorage‐dependent growth are the key steps involved in theL monocytogenessusceptibility phen

ISSN:0248-4900    

DOI:10.1111/j.1768-322X.1995.tb00942.x

出版商:Blackwell Publishing Ltd    

年代:1995

数据来源: WILEY

摘要:

Summary—In the present study we report the occurrence of chaperonins, cpn 10 and cpn60, in Chromatium vinosum and rat hepatocytes, using specific polyclonal antibodies in conjunction with the protein A‐gold immunocytochemical technique. As demonstrated by quantitative evaluations, the immunolabeling for cpn10 and cpn60 in C vinosum cells was associated primarily with the bacterial cell envelope. In rat liver homogenates, Western immunoblotting analysis has shown that antibodies to cpn10 from C vinosum recognize an unique 25‐kDa protein that remains to be further characterized. On the other hand, the antibody to cpn60 from C vinosum revealed the presence of a 60‐kDa protein in the rat liver homogenates. Immunofluorescence on rat liver tissue revealed an intracellular granular labeling for both chaperonins. On the other hand, using the post‐embedding immunoelectron microscopy technique cpn10 and cpn60 were localized specifically in liver mitochondria and peroxisomes. Interestingly, further analysis of the labeling distribution confirmed the association of both proteins with the mitochondrial inner membrane whereas in the peroxisomes the chaperonins appeared to be located in the matrix, away from the limiting peroxisomal membrane. The colocalization of both chaperonins suggests that, as in other bacteria as well as eukaryotic cells, they may act in tandem for the proper folding of particular

ISSN:0248-4900    

DOI:10.1111/j.1768-322X.1995.tb00943.x

出版商:Blackwell Publishing Ltd    

年代:1995

数据来源: WILEY

摘要:

Summary—A human autoimmune serum is used to characterize a protein which is located within a dot‐like structure of the interphase nucleus. The dots are located in the nucleoplasma outside the nucleoli. The dot‐like structure could be observed on a variety of human substrates (HEp‐2, HepG2, HeLa, Molt‐4, WI‐38 cell lines, peripheral blood lymphocytes), mouse cell lines and tissues (3T3, L929, spleen sections), rat sections, marsupial PtK2 cells and hamster cell lines. The range of the number of dots and their size differed between cells and cell lines and varied between 1 and 24. The antigen could be identified as a 53 kDa protein with a pI of 8.7 and was named NDP53. Digestion experiments suggested that the protein is not associated with DNA or RNA, but is associated with the nuclear matrix. Immunelectron microscopy using ultra‐thin sections revealed filamentous structures with a diameter of 0.1 to 0.6 μm. The antigen colocalizes with Sp100, PML and NDP55, which are part of a multiprotein complex known as PML oncogenic domain (POD), nuclear bodies, Kr

ISSN:0248-4900    

DOI:10.1111/j.1768-322X.1995.tb00944.x

出版商:Blackwell Publishing Ltd    

年代:1995

数据来源: WILEY

摘要:

Summary—A profound interrelationship between cytokines and invertebrate (molluses) immune responses has been reported. Different cytokines (IL‐1α, IL‐2 and TNF‐α) significantly stimulate molluscan hemocyte motility, increasing phagocytic activity and provoking the induction of nitric oxide synthase. As far as cell motility is concerned, the response to different cytokines varied between species. These and other recently reported findings (Ottavianiet al(1994)FEBS Lett351, 19–21; Ottavianiet al(1995)Biochem Biophys Res Commun207, 288–292) suggest that cytokines are important, ancestral, and functionally conserved molecules, which have also maintained their pleiotropicity, redundancy in the mode of action, and high promiscuity of their receptors dur

ISSN:0248-4900    

DOI:10.1111/j.1768-322X.1995.tb00945.x

出版商:Blackwell Publishing Ltd    

年代:1995

数据来源: WILEY

摘要:

Summary—Histological and ultrastructural studies have been undertaken in order to identify the particularities and, if possible, some mechanisms involved in regeneration from protoplasts in the eggplantSolanum melongena. The most important result consists of a peculiar and concomitant association of meristematic and differentiated features only in cells from which regenerative processes originat

ISSN:0248-4900    

DOI:10.1111/j.1768-322X.1995.tb00946.x

出版商:Blackwell Publishing Ltd    

年代:1995

数据来源: WILEY