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PubMed1995

Peritrophic envelopes of Lutzomyia spinicrassa (Diptera: Psychodidae).

The secretion, morphology, and chemical composition of the peritrophic envelope were studied in the phlebotomine sand fly, Lutzomyia spinicrassa Morales, OsornoMesa, Osorno & Hoyos, a suspected vector of Leishmania braziliensis in Colombia and Venezuela. Viewed under light microscopy, the envelope matured rapidly and could be dissected from the blood bolus as early as 12 h and until 36 h after feeding; subsequently it began to degrade. The envelope was initially a closed sac around the blood meal, but opened posteriorly in most flies by 6 h. The posterior opening may facilitate the migration and establishment of Le. braziliensis in the hindgut. Secretion of envelope precursors was from the entire midgut epithelium. Electron microscopy revealed that electron-dense precursor material (possibly chitin) was present, bathing the microvilli during the first 12 h after blood feeding. This secretion appeared to originate from the bases of the microvilli. From 1 to 36 h, an electron-lucid precursor material (possibly protein) was secreted from the entire length of microvilli and from their bases. Both precursors appeared to be formed at the epithelial surface, not associated with secretory vesicles. The envelope developed rapidly from precursor material, and by 6 h a defined electron-lucid structure was present above the microvilli. Most mature envelopes (12-36 h) were 0.5-2.1 microns thick, multilayered, wholly electron-lucid, and composed of microfibrils and granules. Electron-dense components were seen in some envelopes at 24-36 h. An anterior hyaline plug was present from 12 to 36 h. Envelopes were composed of chitin, protein, and glycoprotein, based on chemical and histochemical tests. The likely presence of several amino acids (lysine, aspartic acid, and glutamic acid) that may cross-link chitin and protein was demonstrated by a positive ninhydrin-Schiff test. This study constitutes the first ultrastructural investigation of peritrophic envelope development by a New World sand fly.

Animals↗
PubMed1998

Mode of action of lufenuron on larval cat fleas (Siphonaptera: Pulicidae).

Adult cat fleas, Ctenocephalides felis (Bouché), were fed suboptimal in vitro concentrations of lufenuron in blood to allow hatching of flea larvae for cytological study. At concentrations of 0.125, 0.25, and 0.5 ppm, larval hatch was 64, 15, and 4%, respectively. Larvae hatching from eggs laid by adults fed lufenuron at concentrations of 0.025, 0.08, or 0.125 ppm did not differ significantly from the control. However, many larvae from the 0.08-ppm group and higher concentrations died during the 1st instar. Examination of these larvae revealed that they were dying from desiccation caused by bleeding from microscopic lesions in the cuticle or the inability to complete the molt to the next instar. Electron micrographs showed that lufenuron often disrupted formation of the endocuticle resulting in the deposition of an amorphous mass of randomly oriented chitin microfibrils. Other larvae formed normal endocuticle but were unable to digest the old endocuticle or produce new procuticle after apolysis. Failure of larvae to digest old cuticle or form new cuticle was caused by degeneration of the epidermal cells needed for the synthesis of molting fluid and chitin.

Animals↗
PubMed1999

Effect of lufenuron on chorionic and cuticular structure of unhatched larval Ctenocephalides felis (Siphonaptera: Pulicidae).

When adult cat fleas, Ctenocephalides felis (Bouché), were fed concentrations of < or = 0.08 ppm lufenuron in cattle blood, egg hatch did not differ significantly from the controls. However, as the concentration of lufenuron in blood increased from 0.125 to 1.0 ppm egg hatch decreased to 64 and 2%, respectively. Most of these eggs contained fully developed larvae. Microscopic examination of unhatched larvae, revealed that the cuticle epidermal cells, chorion, and vitelline membrane all were affected by lufenuron treatment. Larvae often produced 2 separate cuticles in response to treatment. The 1st cuticle consisted of an indistinct layer of epicuticle and a procuticle composed of randomly deposited chitin microfibrils. After the 1st layer of procuticle separated from the epidermal cells, a 2nd layer of procuticle was deposited. It was not possible to determine whether the egg tooth was functional during larval hatch. The surface of the egg tooth appeared normal, but the cuticle may have had structural abnormalities similar to those seen in other areas of the exoskeleton. Structural defects appeared to be due to the cytotoxic effects of lufenuron. The epidermal cells of treated larvae showed evidence of disintegration (i.e., the nuclei and mitochondria appeared to be degenerating and the amount of endoplasmic reticulum and other cytoplasmic organelles was decreased). The chorion of lufenuron-treated larvae consisted of an outer layer, middle and inner layers that were thinner and less electron dense than those of controls, and lacked the innermost chorionic layer found in the control larvae. The vitelline membrane also was thinner than that of the controls. Larval hatching was prevented by ruptures in the cuticle, which opened during eclosion resulting in the loss of hemolymph and desiccation of the larva. Evidently, tearing of the cuticle was caused by abnormal formation of the procuticle that was not strong enough to withstand the cuticular expansion and muscular movement of the larva within the egg shell.

Animals↗
PubMed1996

Software and database for the analysis of mutations in the human FBN1 gene.

Fibrillin is the major component of extracellular microfibrils. Mutations in the fibrillin gene on chromosome 15 (FBN1) were described at first in the heritable connective tissue disorder, Marfan syndrome (MFS). More recently, FBN1 has also been shown to harbor mutations related to a spectrum of conditions phenotypically related to MFS and many mutations will have to be accumulated before genotype/phenotype relationships emerge. To facilitate mutational analysis of the FBN1 gene, a software package along with a computerized database (currently listing 63 entries) have been created.

DNA Mutational Analysis↗
PubMed1997

Marfan Database (second edition): software and database for the analysis of mutations in the human FBN1 gene.

Fibrillin is the major component of extracellular microfibrils. Mutations in the fibrillin gene on chromosome 15 (FBN1) were described at first in the heritable connective tissue disorder, Marfan syndrome (MFS). More recently, FBN1 has also been shown to harbor mutations related to a spectrum of conditions phenotypically related to MFS. These mutations are private, essentially missense, generally non-recurrent and widely distributed throughout the gene. To date no clear genotype/phenotype relationship has been observed excepted for the localization of neonatal mutations in a cluster between exons 24 and 32. The second version of the computerized Marfan database contains 89 entries. The software has been modified to accomodate new functions and routines.

Animals↗
PubMed1996

Roles of advanced glycation end-products in the progression of diabetic nephropathy.

Available data indicate that the development of diabetic nephropathy is linked to hyperglycaemia. Glucose reacts nonenzymatically with proteins to form Schiff base and Amadori products. Further incubation of these early products leads to the formation of advanced glycation end-products (AGEs). AGEs seem to play a central role in the progression of diabetic nephropathy. Immunohistochemically, AGEs were also detected in an expanded mesangial matrix, especially in nodular lesions from patients with diabetic nephropathy. AGEs staining was noted in the Bowman's capsule, periglomerular fibrosis in sclerosing glomeruli. In our ultrastructural study of mesangial matrix from patients with diabetic nephropathy by high-resolution scanning electron microscopy after cellular removal, the meshwork structure was evident at higher magnification. In nodular lesions, the loose meshwork structure appeared to be composed of various sized strands, ranging from 6 to 24 nm (mean +/- SD: 11.4 +/- 3.8 nm). The pore sizes were variable, ranging from 4 to 70 nm (mean +/- SD: 23.6 +/- 12.3 nm), and were statistically larger than those of normal controls. As the AGEs are localized most notably in nodular lesions, advanced glycations play a role in the progression of diabetic nephropathy through impairment of the assembly of matrix proteins in vivo. Because type V and type VI collagens are the major components of nodular lesions, increases in these interstitial and fibril or microfibril collagens may contribute to the formation of wider strands in the mesangial matrix of a nodular lesion. As no metalloprotease that is specific for type VI collagen has been identified thus far, AGEs formation might occur preferentially in type VI collagen-rich nodular lesions, which are sites of slow turnover.

Diabetic Nephropathies↗
PubMed1976

Synthesis of elastin. A rapid formation of lysine-derived crosslinks by chick embryo aorta.

Aortas of 13-day-old chick embryo were labeled for 0.5 hr with [14C]lysine and subjected to a serial extraction after chase for 1-24 hr with [12C]lysine. Substantial radioactivity was found in insoluble elastin after 3 hr chase. The effect of beta-amino-propionitrile on labeling with [14C]lysine was also examined. Each fraction was hydrolyzed and applied to a short column on an amino acid analyzer. Radioactivity was found in desmosine and isodesmosine of insoluble elastin as early as 1 hr after the beginning of chase. The radioactivity increased rapidly at 2 hr and very slowly thereafter. A large count, which was separated into five peaks on a long column, was observed in other lysine derivatives at 2 hr and increased steadily up to 24 hr, while the lysine count decreased from 1 : 0.5 to 1 : 6 against lysine derivatives and from 1 : 0.04 to 1 : 0.9 against quarter-desmosine after 24 hr. The oxidation of lysine residues incorporated during the 0.5 hr pulse was almost completed during the first 1 hr of chase, and these oxidized residues were incorporated into crosslinks during the following 1 hr. It is suggested that poorly crosslinked elastin accumulated in the soluble fractions. The presence of crosslinking derived from lysine residues was also indicated in the microfibril fraction.

Amino Acids↗
PubMed1998

Increase in the amount of celA1 protein in tobacco BY-2 cells by a cellulose biosynthesis inhibitor, 2,6-dichlorobenzonitrile.

The biochemical analysis of cellulose biosynthesis by plants has been a difficult problem due to the lack of a reliable assay procedure for cellulose synthase activity. Recently, the celA1 gene was cloned from cotton fiber, and this gene was identified from the rsw1 mutant of Arabidopsis as a catalytic subunit of cellulose synthase (Arioli et al. 1998). The cloning of these genes enables us to obtain specific antibodies against cellulose synthase. A highly specific antibody against celA1 protein was prepared and used to detect the protein from microsomal fraction of tobacco BY-2 cells. The quantity of celA1 protein in microsomal fraction of normal BY-2 cells was under the detection limit, although they contained a large quantity of cellulose. In contrast, cells habituated to 1 microM DCB (a specific inhibitor of cellulose biosynthesis) produced 1/10 of cellulose content of the normal cells, but had much more celA1 protein than the normal cells. The amount of polysaccharides in the EDTA-soluble fraction was relatively increased in habituated cells. The results suggest that celA1 protein is stabilized upon DCB binding and that the crystallization of cellulose microfibrils is inhibited simultaneously.

Amino Acid Sequence↗
PubMed2000

Regulation of root growth by gibberellin in Lemna minor.

Hormonal control of root growth was studied in Lemna minor. Although addition of gibberellic acid (GA3) to the culture medium did not promote the root growth, a gibberellin biosynthesis inhibitor, uniconazole P (Un-P), significantly inhibited root growth. Both length and diameter of roots in Un-P-treated plants were significantly smaller than those in control plants, mainly caused by inhibition of cell division. In epidermal cells, the length was slightly decreased and the width increased by Un-P treatment, indicating inhibition of elongation growth. GA3 completely nullified the inhibition caused by Un-P. Transverse cortical microtubules (CMTs) of epidermal cells in the elongation zone were significantly fragmented by treatment with Un-P, but not by that in the presence of GA3. The cellulose microfibril array in the Un-P-treated cells was more random and more oblique than that in the control cells. These results suggested that root growth in L. minor is regulated by endogenous gibberellin.

Gibberellins↗
PubMed2000

Distribution of expansins in graviresponding maize roots.

To test if expansins, wall loosening proteins that disrupt binding between microfibrils and cell wall matrix, participate in the differential elongation of graviresponding roots, Zea mays L. cv. Merit roots were gravistimulated and used for immunolocalization with anti-expansin. Western blots showed cross-reaction with two proteins of maize, one of the same mass as cucumber expansin (29 kDa), the second slightly larger (32 kDa). Maize roots contained mainly the larger protein, but both were found in coleoptiles. The expansin distribution in cucumber roots and hypocotyls was similar to the distribution in maize. Roots showed stronger expansin signals on the expanding convex side than the concave flank as early as 30 min after gravistimulation. Treatment with brefeldin A, a vesicle transport inhibitor, or the auxin transport inhibitor, naphthylphthalamic acid, showed delayed graviresponse and the appearance of differential staining. Our results indicate that expansins may be transported and secreted to cell walls via vesicles and function in wall expansion.

Gravitropism↗
PubMed2006

Cellulose synthesis is required for deposition of reticulate wall ingrowths in transfer cells.

Despite the recognized physiological importance of transfer cells, little is known about how these specialized cells achieve localized deposition of cell wall material, leading to amplification of plasma membrane surface area and enhanced membrane transport capacity. This study establishes that cellulose synthesis is a key early factor in the construction of 'reticulate' wall ingrowths, an elaborate but common form of localized wall deposition characteristic of most transfer cells. Using field emission scanning electron microscopy, wall ingrowths were first visible in epidermal transfer cells of Faba bean cotyledons as raised 'patches' of disorganized and tangled cellulosic material, and, from these structures, ingrowths emerged via further deposition of wall material. The cellulose biosynthesis inhibitors 2,6-dichlorobenzonitrile and isoxaben both caused dramatic reductions in the number of cells depositing wall ingrowths, altered wall ingrowth morphology and visibly disrupted microfibril structure. The restriction of cellulose deposition to discrete patches suggests a novel mechanism for cellulose synthesis in this circumstance. Overall, these results implicate a central role for cellulose synthesis in reticulate wall ingrowth morphology, especially at the initial stage of ingrowth formation, possibly by providing a template for the self-assembly of wall polymers.

Benzamides↗
PubMed1998

NMR analysis of cbEGF domains gives new insights into the structural consequences of a P1148A substitution in fibrillin-1.

Fibrillin-1 is a modular glycoprotein and a major component of the 10-12 nm microfibrils of the extracellular matrix. Mutations in the fibrillin-1 (FBN 1) gene result in the connective tissue disease the Marfan syndrome (MFS) and related disorders. The calcium binding EGF-like (cbEGF) domain is the predominant structural motif of the protein and >70% of mutations leading to MFS disrupt this domain. A missense mutation which changes a proline to alanine (P1148A) in cbEGF domain 13 has been associated with a number of fibrillin disorders including MFS and Shprintzen-Goldberg syndrome. However, it has also been described as a polymorphism. In this study comparative NMR analyses on wild-type and mutant forms of covalently-linked fibrillin cbEGF domain pairs have been performed to investigate the structural consequences of this substitution. A comparison of the two-dimensional NOESY spectra of the wild-type and mutant forms of cbEGF domains 12 & 13 and cbEGF domains 13 & 14 indicated that the proline to alanine amino acid change does not introduce a significant structural defect into cbEGF domain 13 or the adjacent domains and most likely represents a polymorphism. These results demonstrate how, in the case of a protein with a well defined domain organisation such as fibrillin-1, comparative NMR analyses can be used to substantiate genetic evidence for the polymorphic status of an amino acid.

Alanine↗
PubMed2005

Growth strain in the trunk and branches of Chamaecyparis formosensis and its influence on tree form.

Distributions of growth strains in branches, straight trunks and basal sweeping trunks of Chamaecyparis formosensis Matsum. trees were measured with strain gauges. Microfibril angles (MFAs) of the S2 layer of the cell wall were measured by the iodine deposition method and their relationships with growth strain examined. The magnitude of the compressive stress on the lower side of trunks with a basal sweep was greater than that of the tensile stress at the surface of straight trunks. However, transverse compressive stress was similar around the trunk regardless of whether normal wood or compression wood was present. The released surface growth strains varied with MFA. At MFAs of 20-25 degrees , growth stress changed from tension to compression, and compressive stress increased dramatically in the compression wood region. Branches suffer bending stress due to self-loading. This stress is superimposed on the growth stress. Growth strains on the upper or lower sides of branches were larger than those in the trunks, suggesting that generation of growth stress on the lower sides of branches with extensive compression wood is affected by the gravitational bending stress due to self-loading. We conclude that branch form is affected by the interaction between the bending moment due to self-loading and that due to the asymmetric distribution of growth stress. Growth strain distribution in a branch differed depending on whether the branch was horizontal, upward bending or downward bending.

Biomechanical Phenomena↗
PubMed1989

Structure and function of flexure wood in Abies fraseri.

Wood produced during flexure in one-year-old leaders of Abies fraseri (Pursh) Poir. (Fraser fir) was analyzed anatomically and radio-densitometrically. More xylem cells were produced in stems subjected to flexing than in stems that were not flexed. The lumens of tracheids produced in response to flexure were smaller than the lumens of tracheids in normal wood. This was manifest as an increase in the cell wall area/cell lumen area ratio. Microfibril orientation in flexure-induced wood approached the less extreme values found in compression wood. The growth ring composed of flexure-induced wood also had a greater density than normal wood. Compression wood, as defined by cellular characteristics observed in transverse section, was absent in flexed stems. Detailed analysis of the anatomical structure, wood density and biomechanical properties of flexure-induced wood indicated that it has more in common with compression wood than with normal wood.

Journal Article↗
PubMed1994

Biosynthesis of plant cell wall polysaccharides.

The cell wall is the principal structural element of plant form. Cellulose, long crystals of several dozen glucan chains, forms the microfibrillar foundation of plant cell walls and is synthesized at the plasma membrane. Except for callose, all other noncellulosic components are secreted to the cell surface and form a porous matrix assembled around the cellulose microfibrils. These diverse noncellulosic polysaccharides and proteins are made in the endomembrane system. Many questions about the biosynthesis and modification within the Golgi apparatus and integration of cell components at the cell surface remain unanswered. The lability of synthetic complexes upon isolation is one reason for slow progress. However, with new methods of membrane isolation and analysis of products in vitro, recent advances have been made in purifying active synthases from plasma membrane and Golgi apparatus. Likely synthase polypeptides have been identified by affinity-labeling techniques, but we are just beginning to understand the unique features of the coordinated assembly of complex polysaccharides. Nevertheless, such progress renews hope that the first gene of a synthase for a wall polysaccharide from higher plants is within our grasp.

Cell Membrane↗
PubMed1989

Chronologic aging in black skin.

Histologic studies examining chronologic aging in skin have been confined to white skin. In the present study, we examined the features of sun-protected black skin from individuals 6 weeks to 75 years of age with light and electron microscopy. With age, the dermoepidermal junction became flattened with multiple zones of basal lamina and anchoring fibril reduplication. Microfibrils in the papillary dermis became somewhat more irregularly oriented. Compact elastic fibers showed cystic changes and separation of skeleton fibers with age. The area occupied by the superficial vascular plexus in specimens of equal epidermal surface length decreased from the infant to young adult (21-29 years old) to adult (39-52 years old) age groups, then increased in the aged adult (73-75 years old) age group. With the exception of the vascularity in the aged adult group, the above features are similar to those seen in aging white skin, and suggest that chronologic aging in white and black skin is similar. In addition, there was a decrease in the number of melanocytes with age. Basal keratinocyte melanin granule density increased with age to age 52 and remained dense in the aged adult group, even as the number of melanocytes decreased.

Adult↗
PubMed1998

Juvenile hyaline fibromatosis: ultrastructural study.

Juvenile hyaline fibromatosis is a multisystemic disorder characterized by a triad of cephalic fibrous outgrowths, gingival hyperplasia, and flexion contractures. The aim of this study was to find new ultrastructural features that could be useful for differentiating this entity from other types of fibromatosis. Mucosal lesions processed for light and electron microscopy by routine techniques showed hyperactive-appearing spindle-shaped fibroblasts and dysplastic mesenchymal cells. Dilated rough endoplasmic reticulum, prominent Golgi complexes, and multivesicular bodies as well as single membrane vesicles filled with fibrillogranular material were seen within the cytoplasm of dysplastic mesenchymal cells. Many fibrillogranular vesicles contained smaller vesicles. There were also invaginations of the cell membrane that contained fibrillogranular material similar to that seen in the single membrane vesicles, suggesting engulfment of an extracellular substance. The stroma contained both normal and serrated collagen fibrils, microfibrils, and two types of fibrillogranular material, one of them with a characteristic banding pattern. Our clinical and histopathologic findings resemble those previously described in cases of infantile systemic hyalinosis and juvenile hyaline fibromatosis. So many features of these two conditions overlap that it is difficult not to consider them as parts of a spectrum of the same disorder.

Fibroma↗
PubMed2002

Abnormal hepatic expression of fibrillin-1 in children with cholestasis.

Fibrillin-1, one of the main constituents of microfibrils, is present in normal adult liver and overexpressed in fibrotic area around cirrhotic nodules and hepatocellular carcinoma. In this work fibrillin-1 expression was studied by immunohistochemistry in liver samples from children with various cholestatic diseases corresponding to paucity of intrahepatic bile ducts, biliary atresia, congenital hepatic fibrosis, Byler's disease, mitochondrial cytopathy, sclerosing cholangitis, or choledochal cyst. As controls, histologically normal liver samples were used. In control liver, as in adult, fibrillin-1 was expressed in vessel walls, sinusoids, and portal connective tissue, particularly at the interface with the limiting hepatocytic plate and close to the basement membrane of bile ducts. In paucity of intrahepatic bile ducts without fibrosis, the fibrillin-1 distribution was similar to controls. In cholestatic diseases associated with severe fibrosis, such as biliary atresia, congenital hepatic fibrosis, Byler's disease, mitochondrial cytopathy, or sclerosing cholangitis, an enhanced deposition of fibrillin-1 was observed in portal connective tissue and fibrous septa. The strong fibrillin-1 expression close to the basement membrane of biliary structures was lost in cholestatic diseases, except biliary atresia. Finally, in normal and pathologic tissues, fibrillin-1 was co-localized with its putative receptor alphaVbeta3 in sinusoids but not around biliary structures.

Actins↗