A blood-borne PDGF/VEGF-like ligand initiates wound-induced epidermal cell migration in Drosophila larvae

Wound healing is a conserved survival response whose function is to restore the integrity of the tissue after physical trauma. Despite numerous studies in the wound healing field, the signals and pathways that orchestrate and control the wound healing program are still not entirely known. To identify additional signals and pathways that regulate epidermal wound repair in Drosophila larvae, we performed a pilot in vivo RNAi screen using a live reporter for epidermal morphology and a wounding assay. From our pilot screen we identified Pvr, the Drosophila homolog of the vertebrate PDGF/VEGF receptors, and six other genes as epidermal wound healing genes. Morphological analysis of wound-edge cells lacking Pvr or the Drosophila Jun N-terminal Kinase (JNK), previously implicated in larval wound closure, suggest that Pvr signaling leads to cell process extension into the wound site while JNK mediates transient dedifferentiation of wound-edge epidermal cells. Furthermore, we found that one of the three known Pvr ligands, Pvf1, is also required for epidermal wound closure. Through tissue-specific knock down and rescue experiments, we propose a model in which epidermally-produced Pvf1 may be sequestered into the hemolymph (blood) and that tissue damage locally exposes blood-borne Pvf1 to Pvr receptors on epidermal cells at the wound edge, thus initiating epidermal cell process extension and migration into the wound gap. Together, our data suggest that the Pvr and JNK signaling pathways act in parallel to control different aspects of wound closure and that PDGF/VEGF ligands and receptors may have a conserved autocrine role in epidermal wound closure. ^

Mechanism in the Sequential Control of Cell Morphology and S Phase Entry by Epidermal Growth Factor Involves Distinct MEK/ERK Activations

Cell shape plays a role in cell growth, differentiation, and death. Herein, we used the hepatocyte, a normal, highly differentiated cell characterized by a long G1 phase, to understand the mechanisms that link cell shape to growth. First, evidence was provided that the mitogen-activated protein kinase kinase (MEK)/extracellular signal-regulated kinase (ERK) cascade is a key transduction pathway controlling the hepatocyte morphology. MEK2/ERK2 activation in early G1 phase did not lead to cell proliferation but induced cell shape spreading and demonstration was provided that this MAPK-dependent spreading was required for reaching G1/S transition and DNA replication. Moreover, epidermal growth factor (EGF) was found to control this morphogenic signal in addition to its mitogenic effect. Thus, blockade of cell spreading by cytochalasin D or PD98059 treatment resulted in inhibition of EGF-dependent DNA replication. Our data led us to assess the first third of G1, is exclusively devoted to the growth factor-dependent morphogenic events, whereas the mitogenic signal occured at only approximately mid-G1 phase. Moreover, these two growth factor-related sequential signaling events involved successively activation of MEK2-ERK2 and then MEK1/2-ERK1/2 isoforms. In addition, we demonstrated that inhibition of extracellular matrix receptor, such as integrin β1 subunit, leads to cell arrest in G1, whereas EGF was found to up-regulated integrin β1 and fibronectin in a MEK-ERK–dependent manner. This process in relation to cytoskeletal reorganization could induce hepatocyte spreading, making them permissive for DNA replication. Our results provide new insight into the mechanisms by which a growth factor can temporally control dual morphogenic and mitogenic signals during the G1 phase.

Expression of Potato virus Y cytoplasmic inclusion protein in tobacco results in disorganization of parenchyma cells, distortion of epidermal cells, and induces mitochondrial and chloroplast abnormalities, formation of membrane whorls and atypical lipid accumulation

Infection of plant cells by potyviruses induces the formation of cytoplasmic inclusions ranging in size from 200 to 1000 nm. To determine if the ability to form these ordered, insoluble structures is intrinsic to the potyviral cytoplasmic inclusion protein, we have expressed the cytoplasmic inclusion protein from Potato virus Y in tobacco under the control of the chrysanthemum ribulose-1,5-bisphosphate carboxylase small subunit promoter, a highly active, green tissue promoter. No cytoplasmic inclusions were observed in the leaves of transgenic tobacco using transmission electron microscopy, despite being able to clearly visualize these inclusions in Potato virus Y infected tobacco leaves under the same conditions. However, we did observe a wide range of tissue and sub-cellular abnormalities associated with the expression of the Potato virus Y cytoplasmic inclusion protein. These changes included the disruption of normal cell morphology and organization in leaves, mitochondrial and chloroplast internal reorganization, and the formation of atypical lipid accumulations. Despite these significant structural changes, however, transgenic tobacco plants were viable and the results are discussed in the context of potyviral cytoplasmic inclusion protein function.

The role of a kinesin, KLP-20, on neural development and epidermal morphogenesis in Caenorhabditis elegans

The heterotrimeric kinesin-II motor in Caenorhabditis elegans consists of KLP-20, KLP-11, and KAP-1 subunits and broadly functions in cellular transport for the development of biological structures including cilia and axons. The results of this paper support the ubiquitous and necessary role kinesin-II motors have in development, particularly the KLP-20 microtubule-associating subunit. Mutations in klp-20 result in a variable abnormal (vab) phenotype characterized by observable epidermal defects, although the role of this gene in development and the mechanism by which the vab phenotype is produced is largely unknown. The vab phenotype is highly penetrant in the first larval stage (L1) of C. elegans, which supports that klp-20 functions in early development. Ciliated amphid sensory neurons can be stained with a fluorescent dye, DiI, to simultaneously test cilia structure and function, as well as the morphology of the amphid sensory organ. Reduced dye uptake in klp-20 mutant L1s suggests that the microtubule-based cilia are under-developed as a result of defective kinesin-II function. Consistent observations of the PLM mechanosensory neuron using the zdIs5 reporter suggest that klp-20 has an essential role in neuron development, as mutations to klp-20 result in under-developed PLM axons. Qualitative observations suggest there may be an interaction between the development of the overlying epidermis and the underlying nervous system, as a more severe vab phenotype is observed simultaneously with reduced dye uptake, and hence amphid sensory cilia under-development. Furthermore, a more severe vab phenotype manifested as large bumps on the posterior epidermis appears to be spatially correlated with PLM defects. The results presented and discussed in this paper suggest that KLP-20 has a necessary role in neurodevelopment and epidermal morphogenesis in C. elegans during embryogenesis.

Langerhans cell (LC) proliferation mediates neonatal development, homeostasis, and inflammation-associated expansion of the epidermal LC network.

Most tissues develop from stem cells and precursors that undergo differentiation as their proliferative potential decreases. Mature differentiated cells rarely proliferate and are replaced at the end of their life by new cells derived from precursors. Langerhans cells (LCs) of the epidermis, although of myeloid origin, were shown to renew in tissues independently from the bone marrow, suggesting the existence of a dermal or epidermal progenitor. We investigated the mechanisms involved in LC development and homeostasis. We observed that a single wave of LC precursors was recruited in the epidermis of mice around embryonic day 18 and acquired a dendritic morphology, major histocompatibility complex II, CD11c, and langerin expression immediately after birth. Langerin+ cells then undergo a massive burst of proliferation between postnatal day 2 (P2) and P7, expanding their numbers by 10–20-fold. After the first week of life, we observed low-level proliferation of langerin+ cells within the epidermis. However, in a mouse model of atopic dermatitis (AD), a keratinocyte signal triggered increased epidermal LC proliferation. Similar findings were observed in epidermis from human patients with AD. Therefore, proliferation of differentiated resident cells represents an alternative pathway for development in the newborn, homeostasis, and expansion in adults of selected myeloid cell populations such as LCs. This mechanism may be relevant in locations where leukocyte trafficking is limited.

Expression of AmphiCoe, an amphioxus COE/EBF gene, in the developing central nervous system and epidermal sensory neurons

The COE/EBF gene family marks a subset of prospective neurons in the vertebrate central and peripheral. nervous system; including neurons deriving from some ectodermal placodes. Since placodes are often considered unique to vertebrates, we have characterised an amphioxus COE/EBF gene with the aim of using it as a marker to examine the timing and location of peripheral neuron differentiation. A single COE/EBF family member, AmphiCoe, was isolated from the amphioxus Branchiostoma floridae: AmphiCoe lies basal to the vertebrate COE/EBF genes in molecular phylogenetic analysis, suggesting that the duplications that formed the vertebrate COE/EBF family were specific to the vertebrate lineage. AmphiCoe is expressed in the central nervous system and in a small number of scattered ectodermal cells on the flanks of neurulae stage embryos. These cells become at least largely recessed beneath the ectoderm. Scanning electron microscopy was used to examine embryos in which the ectoderm had been partially peeled away. This revealed that these cells have neuronal morphology, and we infer that they are the precursors of epidermal primary sensory neurons. These characters lead us to suggest that differentiation of some ectodermal cells into sensory neurons with a tendency to sink beneath the embryonic surface represents a primitive feature that has become incorporated into placodes during vertebrate evolution. (C) 2004 Wiley-Liss, Inc.

Morphology and anatomy of inflorescence and inflorescence axis in Paepalanthus sect. Diphyomene Ruhland (Eriocaulaceae, Poales) and its taxonomic implications

Paepalanthus sect. Diphyomene has inflorescences arranged in umbels. The underlying bauplan seems however to be more complex and composed of several distinct subunits. Despite appearing superficially very similar, the morphology and anatomy of the inflorescences can supply useful information for the understanding of the phylogeny and taxonomy of the group. Inflorescences of Paepalanthus erectifolius, Paepalanthus flaccidus, Paepalanthus giganteus, and Paepalanthus polycladus were analyzed in regard to branching pattern and anatomy. In P. erectifolius, P. giganteus and P. polycladus the structure is a tribotryum, with terminal dibotryum, and with pherophylls bearing lateral dibotrya. In P. flaccidus, the inflorescence is a pleiobotryum, with terminal subunit, and without pherophylls. Secondary inflorescences may occur in all species without regular pattern. Especially when grown in sites without a pronounced seasonality, the distinction between enrichment zone (part of the same inflorescence) and new inflorescences may be obscured. The main anatomical features supplying diagnostic and phylogenetic information are as follows: (a) in the elongated axis, the thickness of the epidermal cell walls and the cortex size; (b) in the bracts, the quantity of parenchyma cells (c) in the scapes, the shape and the presence of a pith tissue. Therefore, P. sect. Diphyomene can be divided in two groups; group A is represented by P. erectifolius, P. giganteus and P. polycladus, and group B is represented by P. flaccidus. The differentiation is based in both, inflorescence structure and anatomy. Group A presents a life cycle and anatomical features similar to species of Actinocephalus. Molecular trees also point that these two groups are closely related. However, inflorescence morphology and blooming sequence are different. Species of group B present an inflorescence structure and anatomical features shared with many genera and species in Eriocaulaceae. The available molecular and morphology based phylogenies still do not allow a precise allocation of the group in the bulk of basal species of Paepalanthus collocated in P. sect. Variabiles. The characters described and used here supply however important information towards this goal. (C) 2009 Elsevier GmbH. All rights reserved.

Morphology and anatomy of inflorescence and inflorescence axis in Paepalanthus sect. Diphyomene Ruhland (Eriocaulaceae, Poales) and its taxonomic implications

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Morphology and development of anthers and ovules in Croton and Astraea (Euphorbiaceae)

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Morphology and anatomy in Heliconia angusta Vell. and H. velloziana L. Emygd. (Zingiberales: Heliconiaceae) from the Atlantic forest of southeastern Brazil

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

First record of petrified Permian pecopterids from the Parana Basin, Brazil (Corumbatai Formation, Passa Dois Group, northeastern State of São Paulo): Morphology, anatomy and paleoecological implications

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Changes in nuclear morphology and chromatin texture of basal keratinocytes in melasma

BackgroundThe pathogenesis of melasma and the role of keratinocytes in disease development and maintenance are not completely understood. Dermal abnormalities, the expression of inflammatory mediators, growth factors, epithelial expression of melanocortin and sexual hormones receptors suggest that not only melanocytes, but entire epidermal melanin unit is involved in melasma physiopathology.ObjectivesTo compare nuclear morphological features and chromatin texture between basal keratinocytes in facial melasma and adjacent normal skin.MethodsWe took facial skin biopsies (2mm melasma and adjacent normal skin) from women processed for haematoxylin and eosin. Thirty non-overlapping basal keratinocyte nuclei were segmented and descriptors of area, highest diameter, perimeter, circularity, pixel intensity, profilometric index (Ra) and fractal dimension were extracted using ImageJ software.ResultsBasal keratinocyte nuclei from facial melasma epidermis displayed larger size, irregular shape, hyperpigmentation and chromatin heterogeneity by fractal dimension than perilesional skin.ConclusionBasal keratinocytes from facial melasma display changes in nuclear form and chromatin texture, suggesting that the phenotype differences between melasma and adjacent facial skin can result from complete epidermal melanin unit alterations, not just hypertrophic melanocytes.

Analyses of pp60$\sp{{\rm c}-src}$ and epidermal growth factor receptor protein tyrosine kinases in a human colon adenocarcinoma cell line following induction of goblet cell-like characteristics by tumor necrosis factor-alpha

Regulation of colonic epithelial cell proliferation and differentiation remains poorly understood due to the inability to design a model system which recapitulates these processes. Currently, properties of "differentiation" are studied in colon adenocarcinoma cell lines which can be induced to express some, but not all of the phenotypes of normal cells. In this thesis, the DiFi human colon adenocarcinoma cell line is utilized as an in vitro model system in which to study mucin production. In response to treatment with tumor necrosis factor-alpha, DiFi cells acquire some properties of mucin-producing goblet cells including altered morphology, increased reactivity to wheat germ agglutinin, and increased mucin production as determined by RNA expression as well as reactivity with the MUC-1 antibodies, HMFG-1 and SM-3. Thus, TNF-treated DiFi cells represent one of the few in vitro systems in which mucin expression can be induced.^ DiFi cells express an activated pp60$\sp{{\rm c}-src},$ as do most colon adenocarcinomas and derived cell lines, as well as an amplified epidermal growth factor (EGF) receptor. To assess potential changes in these enzymes during induction of differentiation characteristics, potential changes in the levels and activities of these enzymes were examined. For pp60$\sp{{\rm c}-src},$ no changes were observed in protein levels, specific activity of the kinase, cellular localization, or phosphorylation pattern as determined by Staphylococcus aureus V8 protease partial proteolytic mapping after induction of goblet cell-like phenotypic changes. These results suggest that pp60$\sp{{\rm c}-src}$ is regulated differentially in goblet cells than in absorptive cells, as down-modulation of pp60$\sp{{\rm c}-src}$ kinase occurs in the latter. Therefore, effects on pp60$\sp{{\rm c}-src}$ may be critical in colon regulation, and may be important in generating the various colonic epithelial cell types.^ In contrast to pp60$\sp{{\rm c}-src},$ EGF receptor tyrosine kinase activity decreased ($<$5-fold) after TNF treatment and at the time in which morphologic changes were observed. Similar decreases in tyrosine phosphorylation of EGF receptor were observed as assessed by immunoblotting with an anti-phosphotyrosine antibody. In addition, ($\sp{125}$I) -EGF cell surface binding was reduced approximately 3-fold following TNF treatment with a concomitant reduction in receptor affinity ($<$2-fold). These results suggest that modulation of EGF receptor may be important in goblet cell differentiation. In contrast, other published studies have demonstrated that increases in EGF receptor mRNA and in ($\sp{125}$I) -EGF binding accompany differentiation toward the absorptive cell phenotype. Therefore, differential regulation of both EGF receptor and pp60$\sp{{\rm c}-src}$ occur along the goblet cell and absorptive cell differentiation pathways. Thus, my results suggest that TNF-treated DiFi cells represent a unique system in which to study distinct patterns of regulation of pp60$\sp{{\rm c}-src}$ and EGF receptor in colonic cells, and to determine if increased MUC-1 expression is an early event in goblet cell differentiation. ^