Direct demonstration of production of transforming growth factor activity by embryonic chick tissue

Summary The presence of transforming growth factor activity in early chick embryos was directly demonstrated by the ability of limb and tail buds to induce anchorage independent division in NRK 49f cells. Colony number increased with limb bud number and developmental stage. Medium conditioned by tail buds contained some stimulating effect, and strongly promoted the action of other transforming growth factors.     

Molecular and Cellular Basis of Regeneration and Tissue Repair

The Xenopus tadpole is a favourable organism for regeneration research because it is suitable for a wide range of micromanipulative procedures and for a wide range of transgenic methods. Combination of these techniques enables genes to be activated or inhibited at specific times and in specific tissue types to a much higher degree than in any other organism capable of regeneration. Regenerating systems include the tail, the limb buds and the lens. The study of tail regeneration has shown that each tissue type supplies the cells for its own replacement: there is no detectable de-differentiation or metaplasia. Signalling systems needed for regeneration include the BMP and Notch signalling pathways, and perhaps also the Wnt and FGF pathways. The limb buds will regenerate completely at early stages, but not once they are fully differentiated. This provides a good opportunity to study the loss of regenerative ability using transgenic methods.     

General effects of cytosine arabinoside on the embryo of reptiles

Administered to eggs of Reptilia (Lacerta viridis and Anguis fragilis), cytosine-arabinoside induced numerous embryonic malformations, among which microphthalmia and limb deformities. On the other hand this drug delayed the retrogression of the apical ridge of the limb buds in Anguis fragilis.     

Electron microscopic study of the penetration and distribution of somitic cells in the mesoblast of the limb buds of reptiles (Anguis fragilis and Lacerta viridis)]

Based on characteristics of mitochondria and on the amount of lipid inclusions, a distinction between somitic cells and mesoblastic somatopleural cells is possible, at the early stages of the development of the limb bud in Reptiles (Anguis fragilis and Lacerta viridis). The dislocation of the ventral processes of the somites and the localisation of the somitic cells in the mesoblast of the anterior limb buds could be studied.     

Effect of the pentosanpolysulfate SP 54 on the collagen of embryonic limb buds cultured in vitro

Summary After addition of SP 54 to limb buds from 11-day-old mouse embryos in tissue culture, collagen with an altered structure is produced.Durchgeführt mit Unterstützung durch die DFG (SFB 29).     

Effect of sublethal concentrations of sumithion on limb regeneration of fresh water field crab Oziotelphusa senex senex

The initiation and progress of regeneration following the removal of the left 4th walking leg were altered in the crab (Oziotelphusa senex senex) by exposure to sumithion. Depending on the concentration used, sumithion caused a complete inhibition of regeneration, a delay of initiation of limb bud development or a reduction of limb bud growth rate. Crustacean limb regeneration can also be used as a sensitive bioassay for studying the effects of environmental pollutants.     

Morphological and biochemical effects of ethylene on Tulips

Summary Tulip bulbs treated with ethylene at various concentrations, at different stages of development and during various lengths of time, exhibited various morphological abnormalities: open bud, stamen blasting, abnormal number of floral parts, increased number of second-year bulbs. These morphological changes were paralleled by rapid variations in the contents of proteins and RNAs, and a slow, yet noticeable, increase of the DNA content of flower buds.     

Role of growth inhibitors in bud dormancy in virus-infected stem cuttings ofEuphorbia pulcherrima

Summary In virus-infected stem cuttings ofE. pulcherrima, the dormancy in buds is broken in winter possibly because of the low amount of growth inhibitors, abscisic acid and phenols, as compared to healthy stem cuttings.     

Transformation: a tool for studying fungal pathogens of plants

Plant diseases caused by plant pathogenic fungi continuously threaten the sustainability of global crop production. An effective way to study the disease-causing mechanisms of these organisms is to disrupt their genes, in both a targeted and random manner, so as to isolate mutants exhibiting altered virulence. Although a number of techniques have been employed for such an analysis, those based on transformation are by far the most commonly used. In filamentous fungi, the introduction of DNA by transformation typically results in either the heterologous (illegitimate) integration or the homologous integration of the transforming DNA into the target genome. Homologous integration permits a targeted gene disruption by replacing the wild-type allele on the genome with a mutant allele on transforming DNA. This process has been widely used to determine the role of newly isolated fungal genes in pathogenicity. The heterologous integration of transforming DNA causes a random process of gene disruption (insertional mutagenesis) and has led to the isolation of many fungal mutants defective in pathogenicity. A big advantage of insertional mutagenesis over the more traditional chemical or radiation mutagenesis procedures is that the mutated gene is tagged by transforming DNA and can subsequently be cloned using the transforming DNA. The application of various transformation-based techniques for fungal gene manipulation and how they have increased our understanding and appreciation of some of the most serious plant pathogenic fungi are discussed. Received 9 May 2001; received after revision 2 July 2001; accepted 3 July 2001     

Embryonic chick muscle produces an FGF-like activity

Normal and pathological formation of blood vessels is of considerable interest both in terms of basic scientific processes and clinical applications. Angiogenic events in the adult are likely to represent persistence of developmental mechanisms, and embryos are therefore a suitable experimental model for these processes. Among embryonic tissues, muscle is particularly appropriate for investigation, since it is highly vascularised from early stages. There are a number of competing explanations of how this process is controlled. Bioassays offer advantages over conventional molecular localisation techniques, in that they reveal the presence of active processed forms of the molecules under study, rather than non-processed forms, or non-translated meassages. Using these techniques, we report here that embryonic chick muscle, taken from the stages at which blood vessels are forming, produces an angiogenic activity on the chick chorioallantoic membrane (CAM), and transforms NR6 cells in soft agar. Basic fibroblast growth factor (bFGF) is shown to be angiogenic on the CAM in the same way, and also transforms NR6 cells (NR6 cells lack functional epidermal growth factor/transforming growth factor-a receptors, and are believed to respond only to bFGF in this way). Anti-bFGF removes the transforming activity of the embryonic muscle. We conclude that this represents evidence that embryonic chick muscle is producing an FGF-like molecule which is capable of acting as an angiogenic agent at the appropriate times in development.     

Recruitment and retention: factors that affect pericyte migration

Pericytes are critical for vascular morphogenesis and contribute to several pathologies, including cancer development and progression. The mechanisms governing pericyte migration and differentiation are complex and have not been fully established. Current literature suggests that platelet-derived growth factor/platelet-derived growth factor receptor-β, sphingosine 1-phosphate/endothelial differentiation gene-1, angiopoietin-1/tyrosine kinase with immunoglobulin-like and EGF-like domains 2, angiopoietin-2/tyrosine kinase with immunoglobulin-like and EGF-like domains 2, transforming growth factor β/activin receptor-like kinase 1, transforming growth factor β/activin receptor-like kinase 5, Semaphorin-3A/Neuropilin, and matrix metalloproteinase activity regulate the recruitment of pericytes to nascent vessels. Interestingly, many of these pathways are directly affected by secreted protein acidic and rich in cysteine (SPARC). Here, we summarize the function of these factors in pericyte migration and discuss if and how SPARC might influence these activities and thus provide an additional layer of control for the recruitment of vascular support cells. Additionally, the consequences of targeted inhibition of pericytes in tumors and the current understanding of pericyte recruitment in pathological environments are discussed.     

The role of VEGF receptors in angiogenesis; complex partnerships

Vascular endothelial growth factors (VEGFs) regulate blood and lymphatic vessel development and homeostasis but also have profound effects on neural cells. VEGFs are predominantly produced by endothelial, hematopoietic and stromal cells in response to hypoxia and upon stimulation with growth factors such as transforming growth factors, interleukins or platelet-derived growth factor. VEGFs bind to three variants of type III receptor tyrosine kinases, VEGF receptor 1, 2 and 3. Each VEGF isoform binds to a particular subset of these receptors giving rise to the formation of receptor homo- and heterodimers that activate discrete signaling pathways. Signal specificity of VEGF receptors is further modulated upon recruitment of coreceptors, such as neuropilins, heparan sulfate, integrins or cadherins. Here we summarize the knowledge accumulated since the discovery of these proteins more than 20 years ago with the emphasis on the signaling pathways activated by VEGF receptors in endothelial cells during cell migration, growth and differentiation. Received 15 September 2005; received after revision 11 November; accepted 24 November 2005     

Lack of relationship between Langerhans cells,epidermal cell proliferation and epidermal G1 chalone

Summary Mouse tail interscale epidermis contains numerous Langerhans cells, whereas the adjacent scale regions are devoid of these cells. No difference in a) proliferative activity and b) inhibitory effect of the epidermal G1 chalone can be demonstrated in both regions. A direct relationship between Langerhans cells and growth control may be excluded.     

Lack of relationship between Langerhans cells, epidermal cell proliferation and epidermal G1 chalone.

Mouse tail interscale epidermis contains numerous Langerhans cells, whereas the adjacent scale regions are devoid of these cells. No difference in a) proliferative activity and b) inhibitory effect of the epidermal G1 chalone can be demonstrated in both regions. A direct relationship between Langerhans cells and growth control may be excluded.     

Syndecan family of cell surface proteoglycans: Developmentally regulated receptors for extracellular effector molecules

Syndecans are a family of integral membrane proteoglycans with conserved membrane-spanning and intracellular domains but with structurally distinct extracellular domains (ectodomains). They are known to function as heparan sulphate co-receptors in fibroblast growth factor signalling as well as to link cells directly to the extracellular matrix. These and other biological activities of syndecans involve specific interactions of the heparan sulphate side chains of syndecans with cytokines and extracellular matrix proteins. Four different vertebrate syndecans, designated as syndecans 1–4 (or syndecan, fibroglycan, N-syndecan and amphiglycan, respectively), are known. During embryonic development, syndecans have specific and highly regulated expression patterns that are distinct from the expression in adult tissue, suggesting an active role in morphogenetic processes. The developmental expression of syndecans is particularly intense in mesenchymal condensates and at epithelium mesenchyme interfaces, where a number of heparan sulphate-binding cytokines and matrix components are also expressed in a regulated manner, ofter spatially and temporally co-ordinated with the syndecan expression. Recent evidence indicates that the regulation of heparan sulphate fine structure (mainly the number and arrangement of sulphate groups along the polymer) provides a mechanism for the cellular control of syndecan-protein interactions. Furthermore, morphogenetically active cytokines such as fibroblast growth factor-2 and transforming growth factor-β participate in the regulation of syndecan expression and glycosaminoglycan structure. This review discusses the developmental expression and binding functions of syndecans as well as the molecular regulation of specific heparan sulphate-protein interactions.     

Causes of artificially high blood glucose values in experiments with diabetic rats and mice

Unusually high blood sugar values may be caused by the slightest contamination of the blood obtained after puncture of the tail vein by diabetic urine, that has dried onto the tails of the animals. This can be avoided by collecting blood that is allowed to drip from the severed tip of the tail or by carefully cleaning the tail before puncturing the tail vein.     

Causes of artificially high blood glucose values in experiments with diabetic rats and mice

Summary Unusually high blood sugar values may be caused by the slightest contamination of the blood obtained after puncture of the tail vein by diabetic urine, that has dried onto the tails of the animals. This can be avoided by collecting blood that is allowed to drip from the severed tip of the tail or by carefully cleaning the tail before puncturing the tail vein.     

Purification of platelet-derived endothelial cell growth inhibitor and its characterization as transforming growth factor-beta type 1.

In 1986, Brown and Clemmons (Proc. natl Acad. Sci. USA 83 (1986) 3321) showed that platelets contain a substance, platelet-derived growth inhibitor (PDGI), that inhibits in vitro endothelial cell replication. Although platelets are rich in transforming growth factor beta (TGF-beta), PDGI was considered not to be related to TGF-beta, on the basis of its reported properties (extraction from platelets at neutral pH, binding to heparin-Sepharose). However, we purified PDGI to near homogeneity and showed that on the basis of HPLC retention behavior, in vitro growth inhibitory activities with several cell types, receptor binding, and immunoneutralization of growth inhibitory activity with specific anti-TGF-beta type 1 antibodies, PDGI is most probably identical with TGF-beta type 1.     

Thrombospondins: from structure to therapeutics

The thrombospondins (TSPs) are a family of five proteins that are involved in the tissue remodeling that is associated with embryonic development, wound healing, synaptogenesis, and neoplasia. These proteins mediate the interaction of normal and neoplastic cells with the extracellular matrix and surrounding tissue. In the tumor microenvironment, TSP-1 has been shown to suppress tumor growth by inhibiting angiogenesis and by activating transforming growth factor beta. TSP-1 inhibits angiogenesis through direct effects on endothelial cell migration and survival, and through effects on vascular endothelial cell growth factor bioavailability. In addition, TSP-1 may affect tumor cell function through interaction with cell surface receptors and regulation of extracellular proteases. Whereas the role of TSP-1 in the tumor microenvironment is the best characterized, the other TSPs may have similar functions. (Part of a Multi-author Review).     

AMP-activated protein kinase in skeletal muscle: From structure and localization to its role as a master regulator of cellular metabolism

The AMP-activated protein kinase (AMPK) is a metabolite sensing serine/threonine kinase that has been termed the master regulator of cellular energy metabolism due to its numerous roles in the regulation of glucose, lipid, and protein metabolism. In this review, we first summarize the current literature on a number of important aspects of AMPK in skeletal muscle. These include the following: (1) the structural components of the three AMPK subunits (i.e. AMPKα, β, and γ), and their differential localization in response to stimulation in muscle; (2) the biochemical regulation of AMPK by AMP, protein phosphatases, and its three known upstream kinases, LKB1, Ca²⁺/calmodulin-dependent protein kinase kinase (CaMKK), and transforming growth factor-β-activated kinase 1 (TAK1); (3) the pharmacological agents that are currently available for the activation and inhibition of AMPK; (4) the physiological stimuli that activate AMPK in muscle; and (5) the metabolic processes that AMPK regulates in skeletal muscle. Received 04 May 2008; received after revision 14 June 2008; accepted 14 July 2008