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Biomedical subjects

E C Davis

Publications and source records attributed to E C Davis.

35 records · Page 2Linked to original sources
PubMed1996

Unusual presentation of Ewing sarcoma with t(21;22) in a 3-year-old boy.

PURPOSE: We describe a 3-year-old boy with widespread, metastatic Ewing sarcoma and an unusual translocation, involving chromosomes 21 and 22. MATERIALS AND METHODS: Cytogenetic studies were performed on a biopsy of the primary tumor. These included GTG banding and fluorescence in situ hybridization. RESULTS: A balanced translocation between chromosomes 21 and 22 was noted with translocation breakpoints at bands 21q22 and 22q12. CONCLUSIONS: The t(21;22) translocation represents a new cytogenetic abnormality that may be associated with Ewing sarcoma. Its prognostic significance, if any, remains to be determined.

Child, Preschool↗
PubMed1996

Structural sexual dimorphisms in the anteroventral periventricular nucleus of the rat hypothalamus are sensitive to gonadal steroids perinatally, but develop peripubertally.

The volume of the anteroventral periventricular nucleus (AVPv) of the rat hypothalamus is larger in females than in males. A preliminary study from this laboratory found that this sexual dimorphism develops between days 30 and 91. The present study was designed to confirm and extend these findings and to determine the role of endogenous gonadal steroids in the development of the AVPv postnatally. The results indicate that the sexual dimorphism in AVPv volume arises between days 30 and 40 and that the length of the nucleus becomes sexually dimorphic between days 60 and 80. Additionally, both AVPv volume and length increased between days 30 and 80 in females. Castration of male rats on the day of birth sex-reversed AVPv volume in adulthood and AVPv length was sex-reversed by castration of males 5 days after birth; ovariectomy of females at these ages had no effect on either parameter. Moreover, in both males and females, AVPv volume and length were unaffected by gonadectomy at later ages. That the AVPv appears to be influenced by testicular hormones neonatally, but changes structurally around the time of puberty in females, clearly challenges current concepts of sexual differentiation that limit the process to the early postnatal period.

Aging↗
PubMed1995

Elastic fibre assembly: macromolecular interactions.

To investigate the mechanisms behind elastic fibre assembly, we studied the molecular interactions between elastin and microfibrillar components using solid-phase binding assays. Fibrillin 1, purified from tissue using reductive-saline extraction, showed no binding to microfibril-associated glycoprotein (MAGP) or tropoelastin. MAGP, however, was found to bind specifically to tropoelastin in a divalent-cation independent manner. Antibody inhibition studies indicated that the C-terminus of tropoelastin defined the interactive site with MAGP. MAGP and fibrillin were also substrates for transglutaminase, which may provide an important mechanism for stabilizing microfibrillar structure. In other studies we found that a major cross-linking region in elastin is formed through the association of domains encoded by exons 10, 19 and 25 of tropoelastin and that the three chains are joined together by one desmosine and two lysinonorleucine cross-links.

Amino Acid Sequence↗
PubMed1995

Bovine latent transforming growth factor beta 1-binding protein 2: molecular cloning, identification of tissue isoforms, and immunolocalization to elastin-associated microfibrils.

Monoclonal antibodies to fibrillin 1 (MP340), a component of elastin-associated microfibrils, were used to screen cDNA libraries made from bovine nuchal ligament mRNA. One of the selected clones (cL9; 1.2 kb) hybridized on Northern (RNA) blotting with nuchal ligament mRNA to two abundant mRNAs of 9.0 and 7.5 kb, which were clearly distinct from fibrillin mRNA (10 kb). Further library screening and later reverse transcription PCR by the rapid amplification of cDNA ends (RACE) technique resulted in the isolation of additional overlapping cDNAs corresponding to about 6.7 kb of the mRNA. The encoded protein exhibited sequence similarity of around 80% with a recently identified human protein named latent transforming growth factor beta 1 (TGF-beta 1)-binding protein 2 (LTBP-2), indicating that the new protein was bovine LTBP-2. This was confirmed by the specific localization of bovine LTBP-2 cDNA probes to human chromosome 14q24.3, which is the locus of the human LTBP-2 gene. The domain structure of bovine LTBP-2 is very similar to that of the human LTBP-2, containing 20 examples of 6-cysteine epidermal growth factor-like repeats, 16 of which have the consensus sequence for calcium binding, together with 4 examples of 8-cysteine motifs characteristic of fibrillins and LTBP-1. A 4-cysteine sequence which is unique to bovine LTBP-2 and which has similarity to the 8-cysteine motifs was also present. Antibodies raised to two unique bovine LTBP-2 peptides specifically localized in tissue sections to the elastin-associated microfibrils, indicating that LTBP-2 is closely associated with these structures. Immunoblotting experiments identified putative LTBP-2 isoforms as a 260-kDa species released into the medium by cultured elastic tissue cells and as larger 290- and 310-kDa species in tissue extracts. A major proportion of tissue-derived LTBP-2 required treatment with 6 M guanidine for solubilization, indicating that the protein was strongly bound to the microfibrils. Most of the guanidine-solubilized LTBP-2 appeared to be monomeric, indicating that it was not involved in disulfide-bonded aggregation either with itself or with latent TGF-beta. Additional LTBP-2 was resistant to solubilization with 6 M guanidine but was readily extracted with a reductive saline solution. This treatment is relatively specific for solubilization of microfibrillar constituents including fibrillin 1 and microfibril-associated glycoprotein. Therefore, it can be inferred that some LTBP-2 is bound covalently to the microfibrils by reducible disulfide linkages. The evidence suggests that LTBP-2 has a direct role in elastic fiber structure and assembly which may be independent of its growth factor-binding properties. Thus, LTBP-2 appears to share functional characteristics with both LTBP-1 and fibrillins.

Adaptor Proteins, Signal Transducing↗
PubMed1995

A revised critical period for the sexual differentiation of the sexually dimorphic nucleus of the preoptic area in the rat.

The volume of the sexually dimorphic nucleus of the preoptic area (SDN-POA) of the rat is several times larger in males than in females. Several studies have established the importance of gonadal steroids perinatally in the sexual differentiation of the SDN-POA as well as a critical period for the permanent influences of exogenous androgen on the volume of the nucleus. Recent preliminary evidence from this laboratory had suggested, however, that the critical period for the effects of the removal of endogenous gonadal steroids on SDN-POA volume may not match that for the administration of exogenous gonadal steroids. A series of experiments was designed to examine further the effects of the removal of endogenous gonadal steroids on adult SDN-POA volume by castrating male rats at various ages. In spite of a rather clear definition of a postnatal critical period for the effects of exogenous steroid administration, the results of this study indicate that the volume of the SDN-POA is sensitive to the removal of endogenous gonadal steroids for a prolonged period of time, extending through at least day 29 postnatally. The data suggest that there may be multiple critical periods for the sexual differentiation of the SDN-POA and reinforce the concept that these critical periods are distinct from those for other sexually differentiated parameters.

Animals↗
PubMed1995

Elastic lamina growth in the developing mouse aorta.

The growth and development of elastic laminae in the mouse aortic media were investigated by light and electron microscopic autoradiography after a single SC injection of L-[3,4-3H]-valine. Because of the remarkable stability of elastin, radiolabel incorporated into the elastic laminae during early stages of aortic development can be identified in the mature vessel. Light microscopic autoradiographs of aortae from mice injected with radiolabeled valine at 3, 14, or 21 days of postnatal age and sacrificed at 4 months of age showed silver grains evenly distributed around the circumference of the vessel, suggesting uniform elastic lamina growth. Electron microscopic autoradiographs of aortae from mice injected at 3 and 14 days' postnatal age and killed at 4 months of age showed the elastin initially deposited at 3 days to be in the center of the lamina, whereas the elastin deposited at 14 days remained peripherally located. These observations suggest that elastin deposited early in development does not undergo any significant redistribution during growth of the vessel. Because the aorta continues to increase in diameter after the elastic laminae are essentially complete, the fenestrations in the laminae were investigated as possible sites of further expansion of the laminae. In aortae from mice injected at 3 days and sacrificed at 4 days of postnatal age, the edges of the elastic lamina that border on fenestrations showed a large number of silver grains. Regions of the elastic lamina at some distance from the fenestration, however, appeared to be associated with fewer grains. Results from this study not only present unique observations of elastin deposition in developing elastic laminae but also provide evidence that the fenestrations in the elastic laminae may play a role in their continued expansion during later stages of aortic development.

Aging↗
PubMed1995

Fibrillin-1 and fibrillin-2 show temporal and tissue-specific regulation of expression in developing elastic tissues.

The recent characterization of multiple fibrillin genes raises the question of whether each of the fibrillin proteins is a component of elastic fiber microfibrils and whether their expression during development of elastic tissues is consistent with a function associated with elastogenesis. To address these possibilities, the expression of fibrillin-1 and fibrillin-2 was compared with expression of MAGP and tropoelastin in two elastogenic tissues that undergo different developmental programs. For both fibrillins, the greatest increase in expression occurred during the last half of fetal development when elastin production is highest. In fetal bovine nuchal ligament, mRNA levels for fibrillin-1 and fibrillin-2 increased approximately threefold during this period, whereas tropoelastin increased 20-fold. Although the relative increase in expression of both fibrillins was equivalent, the basal level of fibrillin-1 expression was greater than fibrillin-2. In developing bovine aorta, fibrillin mRNA levels again paralleled tropoelastin expression although, compared to ligament, elastin synthesis began at an earlier fetal age in this tissue. Furthermore, the relative increase in aortic fibrillin-2 expression was greater than that for fibrillin-1 and the ratio of fibrillin-2 to fibrillin-1 was higher than in the ligament. In contrast to the fibrillins, MAGP expression in nuchal ligament and aorta remained at a constant high level throughout the fetal period. Indirect immunofluorescent staining and immunoelectron microscopy localized both fibrillins as well as MAGP to elastic fiber microfibrils in these developing tissues. The coordinate upregulation of fibrillin-1 and fibrillin-2 expression with the onset of tropoelastin production is consistent with a role in elastic fiber assembly. Our findings also suggest temporal and tissue-specific regulation for the fibrillins during development.

Age Factors↗
PubMed1994

Conversion to an elastogenic phenotype by fetal hyaline chondrocytes is accompanied by altered expression of elastin-related macromolecules.

Elastic fibers are produced during fetal and neonatal periods, and by maturity elastin expression has ceased. Fetal bovine hyaline chondrocytes acquired an elastogenic phenotype within 24 hr after isolation from the tissue, even though the tissue does not produce elastic fibers or tropoelastin mRNA in vivo. By multiple parameters, hyaline chondrocytes produced elastic fibers that were indistinguishable from those made by elastic chondrocytes derived from fetal elastic ear cartilage. The levels of tropoelastin mRNA, secreted protein, and elastic fiber crosslinks as well as the immunostaining and ultrastructural appearance of elastic fibers produced by cultured hyaline chondrocytes were similar if not identical to those of cultured elastic chondrocytes. We also examined the expression of elastin-associated microfibrillar proteins. In intact hyaline cartilage, we did not detect mRNA for fibrillin 5 mRNA and saw only a relatively weak signal for fibrillin 15 mRNA. These microfibrillar products were expressed with tropoelastin in cultured hyaline chondrocytes as well as in intact elastic cartilage and cultured elastic chondrocytes, suggesting that fibrillin 5 and fibrillin 15 are required for elastic fiber formation. In contrast, the levels of microfibrillar-associated glycoprotein mRNA were decreased in both cell types relative to the high expression seen in vivo. These data indicate that conversion to the elastin phenotype includes induction or modulation of all known components of elastic fibers.

Animals↗
PubMed1994

Structure and expression of fibrillin-2, a novel microfibrillar component preferentially located in elastic matrices.

During the previous cloning of the fibrillin gene (FBN1), we isolated a partial cDNA coding for a fibrillin-like peptide and mapped the corresponding gene (FBN2) to human chromosome 5. (Lee, B., M. Godfrey, E. Vitale, H. Hori, M. G. Mattei, M. Sarfarazi, P. Tsipouras, F. Ramirez, and D. W. Hollister. 1991. Nature [Lond.]. 352:330-334). The study left, however, unresolved whether or not the FBN2 gene product is an extracellular component structurally related to fibrillin. Work presented in this report clarifies this important point. Determination of the entire primary structure of the FBN2 gene product demonstrated that this polypeptide is highly homologous to fibrillin. Immunoelectron microscopy localized both fibrillin proteins to elastin-associated extracellular microfibrils. Finally, immunohistochemistry revealed that the fibrillins co-distribute in elastic and non-elastic connective tissues of the developing embryo, with preferential accumulation of the FBN2 gene product in elastic fiber-rich matrices. These results support the original hypothesis that the fibrillins may have distinct but related functions in the formation and maintenance of extracellular microfibrils. Accordingly, we propose to classify the FBN1 and FBN2 gene products as a new family of extracellular proteins and to name its members fibrillin-1 and fibrillin-2, respectively.

Amino Acid Sequence↗
PubMed1994

Immunolocalization of microfibril and microfibril-associated proteins in the subendothelial matrix of the developing mouse aorta.

In the developing aorta, endothelial cell connecting filaments extend from the abluminal surface of the endothelial cell to the subjacent elastic lamina. The connecting filaments are in alignment with intracellular stress fibers and are oriented parallel to the direction of blood flow. In the present study, the composition of the endothelial cell connecting filaments was investigated by indirect immunogold labeling with antibodies to the microfibril proteins, MP340 (fibrillin) and MAGP, and to fibronectin and heparan sulfate proteoglycan (HSPG). In the subendothelial matrix of both 15-day gestational and 5-day post-natal mouse aortae, the connecting filaments showed moderate immunoreactivity with anti-MP340; however, no significant immunoreaction was seen with anti-MAGP. Anti-fibronectin strongly labeled the connecting filaments and a weak immunoreaction was seen with anti-HSPG. In contrast, the adjacent 'elastin-associated microfibrils' showed a very strong immunoreaction with anti-MP340 and a moderate reaction with anti-MAGP. Little or no reaction was seen with anti-fibronectin or anti-HSPG. The filaments that connect endothelial cells to the subjacent elastic lamina during aortic development are thus microfibrillar in nature and related to elastin-associated microfibrils as evidenced by their positive immunoreaction with anti-MP340. The absence of labeling with anti-MAGP, however, suggests that either these fibrillin-containing filaments do not contain MAGP or that the immunoreactive epitopes are blocked by the proteins that coat the connecting filaments such as fibronectin. These results suggest that microfibrils not in association with elastin may play a role in cell anchorage and, more specifically, in the aorta may be involved in maintaining the structural integrity of the endothelial cell layer during early development of the vessel wall. Furthermore, the absence of immunoreactivity with anti-MAGP on the fibrillin-containing endothelial cell connecting filaments raises the possibility that microfibrils may consist of a family of related filaments rather than a single structural entity.

Actin Cytoskeleton↗
PubMed1993

Stability of elastin in the developing mouse aorta: a quantitative radioautographic study.

Elastic lamina growth during development and the ultimate stability of elastin in the mouse aortic media was investigated by light and electron microscopic radioautography. Following a single subcutaneous injection of L-[3,4-3H]valine at 3 days of age, animals were killed at 9 subsequent time intervals up to 4 months of age. One day after injection, radioautographic silver grains were primarily observed over the elastic laminae; however, silver grains were also seen over the smooth muscle cells and extracellular matrix. By 21 to 28 days of age, the silver grains were almost exclusively located over the elastic laminae. From 28 days to 4 months of age, the distribution of silver grains appeared relatively unchanged. Quantitation of silver grain number/micron2 of elastin showed a steady decrease in the concentration of silver grains associated with the elastic laminae from 4 to 21 days of age. After this time, no significant difference in silver grain concentration was observed. Since the initial decrease in grains/micron2 of elastin corresponds to a period of rapid post-natal growth, the decrease is likely to be a result of dilution of the radiolabel due to new elastin synthesis. With the assumption that little or no significant turnover occurs during this time, a constant growth rate of 4.3% per day was predicted by linear regression analysis. Since no significant difference in the concentration of silver grains was observed from 28 to 118 days of age, no new growth or turnover of elastin can be said to occur during this time period. This is supported by the observation that animals injected with radiolabeled valine at 28 days and 8 months of age showed no significant incorporation of radiolabel into the elastic laminae. The results from this study present the first long-term radioautographic evidence of the stability of aortic elastin and emphasize that initial deposition of elastin and proper assembly of elastic laminae is a critical event in vessel development.

Aging↗
PubMed1993

Endothelial cell connecting filaments anchor endothelial cells to the subjacent elastic lamina in the developing aortic intima of the mouse.

The ultrastructural association of endothelial cells with the subjacent elastic lamina was investigated in the developing mouse aorta by electron microscopy. In the 5-day postnatal aorta, extensive filament bundles extend along the subendothelial matrix connecting the endothelial cells to the underlying elastic lamina. The connecting filaments form lateral associations with the abluminal surface of the endothelial cells in regions of membrane occupied by membrane-associated dense plaques. On the intracellular face of each plaque, the termini of stress fibers penetrate and anchor to the cell membrane in alignment with the extracellular connecting filaments. Both the stress fibers and the connecting filaments are oriented parallel to the longitudinal axis of the vessel. High magnification electron micrographs of individual endothelial cell connecting filaments reveal features similar to those of elastin-associated microfibrils. Each connecting filament consists of a 9-10 nm linear core with an electron-lucent center and peripheral spike-like projections. From the filaments, small thread-like extensions span laterally, linking the filaments into a loose bundle and anchoring them to the endothelial cell membrane and the surface of the elastic lamina. The filaments also appear heavily coated with electron-dense material; often with some degree of periodicity along the filament length. During development, the number of endothelial cell connecting filaments decreases as the elastic lamina expands and the subendothelial matrix is reduced. In the aortic intima of mature mice, the elastic lamina is closely apposed to the abluminal surface of the endothelial cell and no connecting filaments are seen.(ABSTRACT TRUNCATED AT 250 WORDS)

Actin Cytoskeleton↗
PubMed1993

Smooth muscle cell to elastic lamina connections in developing mouse aorta. Role in aortic medial organization.

BACKGROUND: The structural and functional intigration of smooth muscle cells and elastic laminae in the aortic media is not well established. Detailed information concerning normal ultrastructural features of the aortic media will provide a better understanding of the medial changes that occur in vascular diseases such as hypertension and aortic aneurysms. EXPERIMENTAL DESIGN: The ultrastructural development and organization of connections between smooth muscle cells and elastic laminae in the mouse aortic media were studied by light and electron microscopy. RESULTS: Early in development, the smooth muscle cells become linked to the elastic laminae by bundles of microfibrils. These microfibrils become progressively infiltrated with elastin so as to form extensions of elastin from the elastic laminae in the adult media. Each elastin extension spans obliquely from the elastic lamina to the surface of the smooth muscle cell where it attaches in a region of membrane occupied by an intracellular membrane-associated dense plaque. On the cytoplasmic face of the plaque, a contractile filament bundle penetrates and anchors in an orientation similar to that of the extracellular elastin extension. The contractile filament bundle traverses the cell obliquely and anchors in a dense plaque on the opposite side of the cell that is in turn linked to the next elastic lamina by another elastin extension. The extracellular elastin extensions and the intracellular contractile filament bundles thus form a "contractile-elastic unit," a continuous line of structures that links adjacent elastic laminae. The oblique orientation of the contractile-elastic units reverses direction in successive smooth muscle cell layers in a herringbone-like pattern. Thus, tension transmitted to one elastic lamina by the smooth muscle cells on either side results in a uniform force exerted on the elastic lamina in one circumferential direction, that on the adjacent elastic laminae being in the opposite direction. CONCLUSIONS: Results from this study demonstrate the presence of smooth muscle cell to elastic lamina connections that form early in development as contractile-elastic units; basic units of aortic medial ultrastructure. The overall organization of the contractile-elastic units within the aortic media is proposed to provide a means for coordinating contractile and elastic tensions in response to mechanical stresses imposed on the vessel wall.

Aging↗
PubMed1992

Ordered distribution of membrane-associated dense plaques in intact quail gizzard smooth muscle cells revealed by freeze-fracture following treatment with cholesterol probes.

The surface distribution of membrane-associated dense plaques in intact quail gizzard smooth muscle cells was investigated by freeze-fracture. Replicas of fractured smooth muscle cell plasma membrane showed caveola-free regions with few intramembrane particles, interspersed with caveola-populated areas with a higher intramembrane particle density. Electron microscopy of thin sections of quail gizzard smooth muscle revealed the regions free of caveolae to be occupied by membrane-associated dense plaques; anchoring sites for the contractile filaments of the cell. Demarcation between the caveola-populated and caveola-free regions on the relicated intramembrane surface was not clear and thus provided little information concerning the distribution of dense plaque sites. However, treatment of the smooth muscle tissue with the cholesterol-binding agents filipin or tomatin prior to freeze-fracture allowed the dense plaque sites to be easily observed as the sites remained free of the membrane deformations characteristic of these agents. The dense plaque sites consist of caveola-free oval areas juxtaposed in regular bands that traverse the long axis of the cell. The dense plaque sites on the freeze-fracture replica were confirmed by electron microscopy of thin sections of filipin-treated quail gizzard smooth muscle cells, which showed the plasma membrane associated with the dense plaques to be unaffected by the actions of filipin, whereas that of the caveola-populated region was severely deformed. The observations presented in this study provide evidence for a highly ordered distribution of dense plaques at the cell surface of intact quail gizzard smooth muscle cells and thus corroborate existing evidence for an organized substructure of smooth muscle cells.

Animals↗
PubMed1986

Freeze-fracture analysis of intramembrane particles of erythrocytes from normal and dystrophic hamsters.

The intramembrane particles on plasma membranes of erythrocytes of Syrian hamsters afflicted with hereditary muscular dystrophy were compared to those from normal controls by freeze-fracture analysis. The reduced number of particles in both fracture faces of dystrophic erythrocyte plasmalemmae, as compared to that of control red cells, was found to be highly significant (P = .001). Results of this study therefore, support the concept of a generalized membrane defect (abnormality) in muscular dystrophy.

Animals↗
PubMed1983

Reduced sulfur in ashes and slags from the gasification of coals: availability for chemical and microbial oxidation.

This study was initiated to determine if reduced sulfur contained in coal gasifier ash and slag was available for microbial and chemical oxidation because eventual large-quantity landfill disposal of these solid wastes is expected. Continuous application of distilled water to a column containing a high-sulfur-content (4% [wt/wt]) gasifier slag yielded leachates with high sulfate levels (1,300 mg of sulfate liter) and low pH values (4.2). At the end of the experiment, a three-tube most-probable-number analysis indicated that the waste contained 1.3 x 10 thiosulfate-oxidizing bacteria per g. Slag samples obtained aseptically from the column produced sulfate under both abiotic and biotic conditions when incubated in a mineral nutrient solution. Both microbial and chemical sulfate syntheses were greatly stimulated by the addition of thiosulfate to the slag-mineral nutrient solution. These results led to a test of microbial versus chemical sulfur oxidation in ashes and slags from five gasification processes. Sulfate production was measured in sterile (autoclaved) and nonsterile suspensions of the solid wastes in a mineral nutrient solution. These ashes and slags varied in sulfur content from 0.3 to 4.0% (wt/wt). Four of these wastes demonstrated both chemical (2.0 to 27 mug of sulfate g day) and microbial (3.1 to 114 mug of sulfate g day) sulfur oxidation. Obvious relationships between sulfur oxidation rate and either sulfur content or particle size distribution of the wastes were not immediately evident. We conclude that the sulfur contained in all but one waste is available for oxidation to sulfuric acid and that microorganisms play a partial role in this process.

Journal Article↗