The impact of metamorphosis on the cranial osteology of giant salamanders of the genus Dicamptodon

Pacific giant salamanders (Dicamptodon) rank among the largest terrestrial caudates. Their ontogeny produces two distinct morphs—larval‐neotenic and metamorphosed—which differ in many morphological traits. We identified changes that are initiated by metamorphosis (distinguishing transformed from neotenic specimens) and also recognized age‐related changes occurring irrespective of transformation. During metamorphosis, specimens remodel the palate, rearrange the vomerine dentition, expand the maxilla, broaden the cheek, foreshorten the posterior skull table and develop specific serrated suture patterns in the dermal bones. Instead, large larvae grow a robust pterygoid sutured with a fully ossified trapezoidal vomer and a short maxilla. Small larvae are readily distinguished by tooth count, morphology and arrangement from more advanced larvae. Age‐related features, irrespective of metamorphosis, include pedicellate teeth, morphological differentiation of parasphenoid, enlargement of the orbitosphenoid, distal expansion of columella, and loss of coronoid teeth.     

Examination of Duct Physiology in the Human Mammary Gland

BackgroundThe human breast comprise several ductal systems, or lobes, which contain a small amount of fluid containing cells, hormones, proteins and metabolites. The complex physiology of these ducts is likely a contributing factor to the development of breast cancer, especially given that the vast majority of breast cancers begin in a single lobular unit.MethodsWe examined the levels of total protein, progesterone, estradiol, estrone sulfate, dehydroepiandrosterone sulfate, and macrophages in ductal fluid samples obtained from 3 ducts each in 78 women, sampled twice over a 6 month period. Samples were processed for both cytological and molecular analysis. Intraclass correlation coefficients and mixed models were utilized to identify significant data.ResultsWe found that the levels of these ductal fluid components were generally uncorrelated among ducts within a single breast and over time, suggesting that each lobe within the breast has a distinct physiology. However, we also found that estradiol was more correlated in women who were nulliparous or produced nipple aspirate fluid.ConclusionsOur results provide evidence that the microenvironment of any given lobular unit is unique to that individual unit, findings that may provide clues about the initiation and development of ductal carcinomas.     

Osteopenic mice: animal models of the aging skeleton

While our understanding of the developmental biology of the skeleton, like that of virtually every other subject in biology, has been transformed by recent advances in human and mouse genetics, we still know very little, in molecular and genetic terms, about skeletal physiology. Thus, among the many questions that are largely unexplained are the following: why is osteoporosis mainly a women's disease? How is bone mass maintained nearly constant between the end of puberty and the arrest of gonadal functions? Molecular genetics has emerged as a powerful tool to study previously unexplored aspects of the physiology of the skeleton. Among mammals, mice are the most promising animals for this experimental work. This has been previously demonstrated e.g. through the tremendous impact of the different osteopetrotic models on our molecular understanding of osteoclastic bone resorption. Until recently the only way of studying bone loss situations and osteoporosis in mice was by using ovariectomy with all its limitations. Today, however, we have access to more sophisticated osteoporotic mouse-models from four different origins: Transgenic mice (HSV-TK), knock-out mice (OPG), inbred-strains (SAMP6), and through physiological modulation (icv application). These new models have already taught us several important lessons. The first is, that bone remodeling is more than just an autocrine/paracrine process. Multiple experimental evidence has demonstrated that the latter regulation exists, but genetics prove that there is no functional cross-control between resorption and formation. The second lesson is, that remodeling is, at least in part, subject to central regulation. Thus, osteoporosis is partly a central or hypothalamic disease. However, the most dramatic change and the most important advantage we feel is, that today we have models to test a new hypothesis regarding the etiology of osteoporosis before it turns to dogma. Taken together, mouse-studies may lead to a shift in our physiological understanding of skeleton biology and to the emergence of novel paradigms. These, in turn, should help us to devise new treatments for degenerative diseases of the skeleton such as osteoporosis and its associated clinical problems.     

Depth-sensing analysis of cytoskeleton organization based on AFM data

Atomic force microscopy is a common technique used to determine the elastic properties of living cells. It furnishes the relative Young’s modulus, which is typically determined for indentation depths within the range 300–500 nm. Here, we present the results of depth-sensing analysis of the mechanical properties of living fibroblasts measured under physiological conditions. Distributions of the Young’s moduli were obtained for all studied cells and for every cell. The results show that for small indentation depths, histograms of the relative values of the Young’s modulus described the regions rich in the network of actin filaments. For large indentation depths, the overall stiffness of a whole cell was obtained, which was accompanied by a decrease of the modulus value. In conclusion, the results enable us to describe the non-homogeneity of the cell cytoskeleton, particularly, its contribution linked to actin filaments located beneath the cell membrane. Preliminary results showing a potential application to improve the detection of cancerous cells, have been presented for melanoma cell lines.     

East European lowland as an area of long-time interaction between Caucasoid and Mongoloid peoples

Most of the population of Eastern Europe inhabit an area of great anthropological interest, because of the contact between Caucasoid and Mongoloid anthropological types. We have analyzed normal variability in minisatellite and microsatellite loci in some East European population. Different synthetic maps were constructed using reliability theory to evaluate the degree of accuracy. Comparison of the synthetic maps for DNA with classical markers has revealed a high level of correlation. All the data obtained show the diverse influence of both anthropological types in forming the gene pool of the Eastern European peoples.     

Lack of bufadienolides in the skin secretion of red bellied toads, Melanophryniscus spp. (Anura, Bufonidae), from Uruguay

The South-American red bellied toads (Melanophryniscus spp.) belonging to the Bufonidae family contain toxic alkaloids in their skin, predominantly of the pumiliotoxin group. Whole animal methanolic extracts of individual specimens of three species (Melanophryniscus atroluteus, M. devincenzii, and M. montevidensis) were analyzed for the presence of toad specific bufadienolides and indolalkylamines (serotonin derivatives) by HPLC-electrospray (ESI)-MS-TOF. No bufadienolides, but few bufotenines, mainly dehydrobufotenine, were detected in the extracts in variable amounts. The concentration of the dehydrobufotenine in the extracts seems to be species specific. Whereas M. atroluteus and M. montevidensis contain very low or trace amounts, M. devincenzii specimens exhibit high concentrations of this indolalkylamine. In comparison, analysis of extracts from Bufo arenarum (Uruguay) and from B. bufo (Germany) confirmed the presence of bufadienolides as well as of bufotenine derivatives. Tadpoles of both species exhibited a different pattern: extracts from B. arenarum tadpoles contained only dehydrobufotenine, but those from B. bufo tadpoles bufotoxin and two alkylamines. Melanophryniscus toads appear not to be able to compensate the high variability of toxic skin alkaloids by producing defensive bufadienolides.     

Characterization of Conserved Region 2-Deficient Mutants of the Cytomegalovirus Egress Protein pM53

Dominant-negative (DN) mutants are powerful tools for studying essential protein-protein interactions. A systematic genetic screen of the essential murine cytomegalovirus (MCMV) protein pM53 identified the accumulation of inhibitory mutations within conserved region 2 (CR2) and CR4. The strong inhibitory potential of these CR4 mutants is characterized by a particular phenotype. The DN effect of the small insertion mutations in CR2 was too weak to analyze (M. Popa, Z. Ruzsics, M. Lötzerich, L. Dölken, C. Buser, P. Walther, and U. H. Koszinowski, J. Virol. 84:9035–9046, 2010); therefore, the present study describes the construction of M53 alleles lacking CR2 (either completely or partially) and subsequent examination of the DN effect on MCMV replication upon conditional expression. Overexpression of CR2-deficient pM53 inhibited virus production by about 10,000-fold. This was due to interference with capsid export from the nucleus and viral genome cleavage/packaging. In addition, the fate of the nuclear envelopment complex in the presence of DN pM53 overexpression was analyzed. The CR2 mutants were able to bind to pM50, albeit to a lesser extent than the wild-type protein, and relocalized the wild-type nuclear envelope complex in infected cells. Unlike the CR4 DN, the CR2 DN mutants did not affect the stability of pM50.     

A genetic screen identifies genes essential for development of myelinated axons in zebrafish

The myelin sheath insulates axons in the vertebrate nervous system, allowing rapid propagation of action potentials via saltatory conduction. Specialized glial cells, termed Schwann cells in the PNS and oligodendrocytes in the CNS, wrap axons to form myelin, a compacted, multilayered sheath comprising specific proteins and lipids. Disruption of myelinated axons causes human diseases, including multiple sclerosis and Charcot-Marie-Tooth peripheral neuropathies. Despite the progress in identifying human disease genes and other mutations disrupting glial development and myelination, many important unanswered questions remain about the mechanisms that coordinate the development of myelinated axons. To address these questions, we began a genetic dissection of myelination in zebrafish. Here we report a genetic screen that identified 13 mutations, which define 10 genes, disrupting the development of myelinated axons. We present the initial characterization of seven of these mutations, defining six different genes, along with additional characterization of mutations that we have described previously. The different mutations affect the PNS, the CNS, or both, and phenotypic analyses indicate that the genes affect a wide range of steps in glial development, from fate specification through terminal differentiation. The analysis of these mutations will advance our understanding of myelination, and the mutants will serve as models of human diseases of myelin.     

The proneural gene ascl1a is required for endocrine differentiation and cell survival in the zebrafish adenohypophysis

Mammalian basic helix-loop-helix proteins of the achaete-scute family are proneural factors that, in addition to the central nervous system, are required for the differentiation of peripheral neurons and sensory cells, derivatives of the neural crest and placodal ectoderm. Here, in identifying the molecular nature of the pia mutation, we investigate the role of the zebrafish achaete-scute homologue ascl1a during development of the adenohypophysis, an endocrine derivative of the placodal ectoderm. Similar to mutants deficient in Fgf3 signaling from the adjacent ventral diencepahalon, pia mutants display failure of endocrine differentiation of all adenohypophyseal cell types. Shortly after the failed first phase of cell differentiation, the adenohypophysis of pia mutants displays a transient phase of cell death, which affects most, but not all adenohypophyseal cells. Surviving cells form a smaller pituitary rudiment, lack expression of specific adenohypophyseal marker genes (pit1, neurod), while expressing others (lim3, pitx3), and display an ultrastructure reminiscent of precursor cells. During normal development, ascl1a is expressed in the adenohypophysis and the adjacent diencephalon, the source of Fgf3 signals. However, chimera analyses show that ascl1a is required cell-autonomously in adenohypophyseal cells themselves. In fgf3 mutants, adenohypophyseal expression of ascl1a is absent, while implantation of Fgf3-soaked beads into pia mutants enhances ascl1a, but fails to rescue pit1 expression. Together, this suggests that Ascl1a might act downstream of diencephalic Fgf3 signaling to mediate some of the effects of Fgf3 on the developing adenohypophysis.