A fat body derived protein is selectively sulfated in the stick insect ovary by transcytosis through the follicular epithelium

With the onset of vitellogenesis, the follicular epithelium overlying the oocyte in stick insect ovarioles becomes highly polarized and patent by formation of wide intercellular spaces. The aim of the present study was to provide experimental support to the notion that the follicular epithelium in this insect species may be involved in transcytosis. Data demonstrate that the follicular epithelium carries out sulfo-conjugation of a 85 kDa fat body derived protein by allowing it to transit from one cell pole to another. Along the basal end, follicle cells branch into a number of cytoplasmic finger-like projections. At the opposite end facing the oocyte they taper off into lance-head shapes. Different vesicular elements are evident at both these extremities. In vivo exposure to horseradish peroxidase shows that the vesicular elements present along the apical end provide an endocytic entry. In contrast, those present along the basal end are labeled with sodium [³⁵S]-sulfate, suggesting that they may be exocytic vesicles containing a sulfo-conjugated secretory product. In vivo exposure to sodium [³⁵S]-sulfate caused radioactivity to appear over the Golgi apparatus and some nearby vesicles of the follicle cell cytoplasm, including the exocytic vesicles. The intracellular pathway of the follicle cells was also examined by immunogold labeling using a monoclonal antibody raised against a 85 kDa fat body derived protein. Under these conditions, gold particles were consistently detected over the Golgi apparatus and the vesicular elements lying along both poles of the follicle cell membrane. Based on this evidence, it is concluded that follicular cells in stick insect ovarioles are endowed with the ability to undergo transcytosis by providing an endocytic entry along the apical end and by releasing exocytically a sulfo-conjugated 85 kDa protein along the baso-lateral domain of the follicle cell membrane.     

PK 11195 differentially affects cell survival in human wild-type and 18 kDa translocator protein-silenced ADF astrocytoma cells

Gliomas are the most common brain tumours with a poor prognosis due to their aggressiveness and propensity for recurrence. The 18 kDa translocator protein (TSPO) has been demonstrated to be greatly expressed in glioma cells and its over-expression has been correlated with glioma malignance grades. Due to both its high density in tumours and the pro-apoptotic activity of its ligands, TSPO has been suggested as a promising target in gliomas. With the aim to evidence if the TSPO expression level alters glioma cell susceptibility to undergo to cell death, we analysed the effects of the specific TSPO ligand, PK 11195, in human astrocytoma wild-type and TSPO-silenced cell lines. As first step, TSPO was characterised in human astrocytoma cell line (ADF). Our data demonstrated the presence of a single class of TSPO binding sites highly expressed in mitochondria. PK 11195 cell treatment activated an autophagic pathway followed by apoptosis mediated by the modulation of the mitochondrial permeability transition. In TSPO-silenced cells, produced by siRNA technique, a reduced cell proliferation rate and a decreased cell susceptibility to the PK 11195-induced anti-proliferative effect and mitochondrial potential dissipation were demonstrated respect to control cells. In conclusion, for the first time, PK 11195 was demonstrated to differentially affect glioma cell survival in relation to TSPO expression levels. These results encourage the development of specific-cell strategies for the treatment of gliomas, in which TSPO is highly expressed respect to normal cells.     

Two msh/msx-related genes, Djmsh1 and Djmsh2, contribute to the early blastema growth during planarian head regeneration

Regeneration in planarians is an intriguing phenomenon, based on the presence of pluripotent stem cells, known as neoblasts. Following amputation, these cells activate mitotic divisions, migrate distally and undergo differentiation, giving rise to the regeneration blastema. We have identified two msh/msx-related genes, Djmsh1 and Djmsh2, which are expressed in distinct cell populations of the planarian Dugesia japonica and activated, with different patterns, during head regeneration. We demonstrate that RNA interference of Djmsh1 or Djmsh2 generates a delay in the growth of cephalic blastema, interfering with the dynamics of mitoses during its initial formation. Our data also reveal that the activity of the two planarian msh genes is required to regulate Djbmp expression during head regeneration. This study identifies, for the first time, a functional association between muscle segment homeobox (MSH) homeoproteins and BMP signaling during stem cell-based regeneration of the planarian head and provides a functional analysis of how msh genes may regulate in vivo the regenerative response of planarian stem cells.     

Adult stem cell plasticity: Neoblast repopulation in non-lethally irradiated planarians

Planarians are a model system for studying adult stem cells, as they possess the neoblasts, a population of pluripotent adult stem cells able to give rise to both somatic and germ cells. Although over the last years several efforts have been made to shed light on neoblast biology, only recent evidence indicate that this population of cells is heterogeneous. In this study we irradiated planarians with different non-lethal X-ray doses (1-5 Gy) and we identified subpopulations of neoblasts with diverse levels of tolerance to X-rays. We demonstrated that a dramatic reduction of neoblasts occurred soon after non-lethal irradiations and that de-novo proliferation of some radioresistant cells re-established the primary neoblast number. In particular, a strong proliferation activity occurred at the ventral side of irradiated animals close to the nervous system. The produced cells migrated towards the dorsal parenchyma and, together with some dorsal radioresistant cells, reconstituted the entire neoblast population demonstrating the extreme plasticity of this adult stem cell system.     

Genetic network of the eye in Platyhelminthes: expression and functional analysis of some players during planarian regeneration

Planarians are the free-living members (order Tricladida) of the phylum Platyhelminthes. They are triploblastic, acoelomate, unsegmented and located at the base of the Lophotrochozoa clade. Besides their huge regenerative capacity, planarians have simple eyes, considered similar to the prototypic eye suggested by Charles Darwin in his book 'On the Origin of Species'. The conserved genetic network that determines the initial steps of eye development across metazoans supports a monophyletic origin of the various eye types present in the animal kingdom. Here we summarise the pattern of expression of certain genes involved in the eye network that have been isolated in planarians, such as Otx, Pax-6, Six, Rax and opsin. We describe the effects of RNA interference-mediated loss of function on eye regeneration. Finally, we discuss the relevance of these findings for the evolution of the eye gene network.