Prosaposin Facilitates Sciatic Nerve Regeneration In Vivo

Abstract: Prosaposin, a multifunctional protein, is the precursor of saposins, which activate sphingolipid hydrolases. In addition to acting as a precursor for saposins, prosaposin has been shown to rescue hippocampal CA1 neurons from lethal ischemic damage in vivo and to promote neurite extension of neuroblastoma cells in vitro. Here we show that prosaposin, when added to a collagen-filled nerve guide after sciatic nerve transection in guinea pigs, increased dramatically the number of regenerating nerve fibers within the guide. To identify the target neurons of prosaposin during peripheral nerve regeneration, we determined the degree of atrophy and chromatolysis of neurons in the spinal anterior horn and dorsal root ganglia on the prosaposin-treated and untreated side. The effect of prosaposin on large spinal neurons and small neurons of the dorsal root ganglion was more conspicuous. Subsequent immunohistochemistry demonstrated that the atrophy of cholinergic large neurons in the anterior horn is prevented to significant extent by prosaposin treatment. These findings suggest that prosaposin promotes peripheral nerve regeneration by acting on α-motor neurons in the anterior horn and on small sensory neurons in the dorsal root ganglion. The present study raises the possibility of using prosaposin as a tool for the treatment of peripheral nerve injuries.     

Localized delivery of epidermal growth factor improves the survival of transplanted hepatocytes

Hepatocyte transplantation may provide a new approach for treating a variety of liver diseases if a sufficient number of the transplanted cells survive over an extended time period. In this report, we describe a technique to deliver growth factors to transplanted hepatocytes to improve their engraftment. Epidermal growth factor (EGF) was incorporated (0.11%) into microspheres (19 +/- 12 mum) fabricated from a copolymer of lactic and glycolic acid using a double emulsion technique. The incorporated EGF was steadily released over 1 month in vitro, and it remained biologically active, as determined by its ability to stimulate DNA synthesis, cell division, and long-term survival of cultured hepatocytes. EGF-containing microspheres were mixed with a suspension of hepatocytes, seeded onto porous sponges, and implanted into the mesentery of two groups of Lewis rats. The first group of animals had their portal vein shunted to the inferior vena cava prior to cell transplantation (portal-caval shunt = PCS), and the second group of animals did not (non-PCS). This surgical procedure improves the survival of transplanted hepatocytes. The engraftment of transplanted hepatocytes in PCS animals was increased two-fold by adding EGF microspheres, as compared to adding control microspheres that contained no growth factors. Devices implanted into non-PCS animals had fewer engrafted hepatocytes than devices implanted into PCS animals, regardless of whether blank or EGF-containing microspheres were added. These results first indicate that it is possible to design systems which can alter the microenvironment of transplanted hepatocytes to improve their engraftment. They also suggest that hepatocyte engraftment is not improved by providing single growth factors unless the correct environment (PCS) is provided for the transplanted cells. (c) 1996 John Wiley & Sons, Inc.     

RecA, Tus protein and constitutive stable DNA replication inEscherichia coli rnhA mutants

Constitutive stable DNA replication (cSDR), which uniquely occurs inEscherichia coli rnhA mutants deficient in ribonuclease HI activity, requires RecA function. TherecA428 mutation, which inactivates the recombinase activity but imparts a constitutive coprotease activity, blocks cSDR inrnhA mutants. The result indicates that the recombinase activity of RecA, which promotes homologous pairing and strand exchange, is essential for cSDR. Despite the requirement for RecA recombinase activity, mutations inrecB, recD, recJ, ruvA andruvC neither inhibit nor stimulate cSDR. It was proposed that the property of RecA essential for homologous pairing and strand exchange is uniquely required for initiation of cSDR inrnhA mutants without involving the homologous recombination process. The possibility that RecA protein is necessary to counteract the action of Tus protein, a contra-helicase which stalls replication forks in theter region of the chromosome, was ruled out because introduction of thetus : :kan mutation, which inactivates Tus protein, did not alleviate the RecA requirement for cSDR.     

Molecular characterization of rgp2, a gene encoding a small GTP-binding protein from rice

Summary We previously reported the isolation of rgp1, a gene from rice, which encodes a ras-related GTP-binding protein, and subsequently showed that the gene induces specific morphological changes in transgenic tobacco plants. Here, we report the isolation and characterization of an rgp1 homologue, rgp2, from rice. The deduced rgp2 protein sequence shows 53% identity with the rice rgp1 protein, but 63% identity with both the marine ray ora3 protein, which is closely associated with synaptic vesicles of neuronal tissue, and the mammalian rab11 protein. Conservation of particular amino acid sequence motifs places rgp2 in the rab/ypt subfamily, which has been implicated in vesicular transport. Northern blot analysis of rgp1 and rgp2 suggests that both genes show relatively high, but differential, levels of expression in leaves, stems and panicles, but low levels in roots. In addition, whereas rgp1 shows maximal expression at a particular stage of plantlet growth, rgp2 is constitutively expressed during the same period. Southern blot analysis suggests that, in addition to rgp1 and rgp2, several other homologues exist in rice and these may constitute a small multigene family.     

Changes in cell surface charges during differentiation of isolated micromeres and mesomeres from sea urchin embryos

Changes in the negative surface charge were observed by cell electrophoresis during the differentiation of micromeres and mesomeres isolated from 16-cell-stage sea urchin embryos. Micromeres and mesomeres were separated by a sucrose density gradient column and were cultured in normal seawater. An isolated micromere developed to a cell aggregate, and, at the mesenchyme-blastula stage of control, the aggregate began to scatter into single cells. These processes are quite similar to those of the primary mesenchyme cells in situ. An isolated mesomere, on the other hand, developed into an ectodermal vesicle. At desired stages of development, the cell aggregates which derived from single blastomeres were dissociated into single cells, and their electrophoretic mobilities were measured. It was found that the electrophoretic mobility of the micromere- and mesomere-derived cells concomitantly increased from the early blastula stage up to the early mesenchyme stage. In contrast with the mesomere-derived cells, however, the micromere-derived cells showed another increase in electrophoretic mobility when the cells began to migrate as primary mesenchyme cells. These results show that a correlation exists between the increase in cell surface negative charge and the migration of the primary mesenchyme cells.     

Morphological and functional peculiarities of mesenchymal cells in the pars tuberalis of the pituitary gland

Summary Following injection of high doses of horseradish peroxidase (HRP), mesenchymal cells distributed in the perisinusoidal space of the pars tuberalis of the hypophysis in cats, rabbits and Japanese quails, sequester the exogenously administrated peroxidase intensively. These cells are designated by the authors as horseradish peroxidase-uptake cells (HRP-uptake cells or HUC). HRP-uptake cells constitute a system of macrophages in the pars tuberalis of mammals and birds, and are located around the hypophysial portal veins. HRP-uptake cells differ in morphological and functional characteristics from similar cells in other parts of hypophysis. They are thought to play a role in the hypothalamic control of adenohypophysial secretion.Supported by grants (No. 144022, 237002) from the Ministry of Education, Science and Culture, Japan     

Morphological studies on neuroglia

Summary The silver-impregnation procedure of Tsujiyama is suitable for demonstration of all three classical types of neuroglial cells; in the present study it was used for electron microscopic identification of neuroglial cells in the brain of the cat. The aim of the present study was 1) to determine impregnated structural correlates of neuroglial cells at the light- and electron-microscopic levels, and 2) to determine whether the method of Tsujiyama is applicable for the electron microscopic identification of the single types of neuroglial cells. Silver deposits were observed over the cytoplasm and processes of astrocytes where numerous glial filaments were present. Oligodendrocytes and microglial cells may be precisely differentiated by use of Tsujiyama's silver impregnation method at the electron microscopic level due to the pattern of silver-deposition in these two basic types of cells. This silver-impregnation method combined with electron microscopy is thus suitable for a precise identification of neuroglial cells; the technique may prove to be very helpful in identification of such categories of neuroglial cells that encompass also the images of cells which cannot be classified by use of the standard methods.Supported by a grant (No. 437002) from the Ministry of Education, Science and Culture, Japan