RESUMO
BACKGROUND: The Quebec platelet disorder (QPD) is inherited and characterized by delayed-onset bleeding following hemostatic challenge. Other characteristics include increased expression and storage of active urokinase-type plasminogen activator (u-PA) in platelets in the setting of normal to increased u-PA in plasma. There is also consumption of platelet plasminogen activator inhibitor-1 and increased generation of plasmin in platelets accompanied by proteolysis of stored alpha-granule proteins, including Factor V. AIMS AND METHODS: Although fibrinolysis has been proposed to contribute to QPD bleeding, the effects of QPD blood and platelets on clot lysis have not been evaluated. We used thromboelastography (TEG), biochemical evaluations of whole blood clot lysis, assessments of clot ultrastructure, and perfusion of blood over preformed fibrin to gain insights into the disturbed hemostasis in the QPD. RESULTS: Thromboelastography was not sensitive to the increased u-PA in QPD blood. However, there was abnormal plasmin generation in QPD whole blood clots, generated at low shear, with biochemical evidence of increased fibrinolysis. The incorporation of QPD platelets into a forming clot led to progressive disruption of fibrin and platelet aggregates unless drugs were added to inhibit plasmin. In whole blood perfusion studies, QPD platelets showed normal adherence to fibrin, but their adhesion was followed by accelerated fibrinolysis. CONCLUSIONS: The QPD is associated with "gain-of-function" abnormalities that increase the lysis of forming or preformed clots. These findings suggest accelerated fibrinolysis is an important contributor to QPD bleeding.
Assuntos
Transtornos Plaquetários/fisiopatologia , Plaquetas/patologia , Adesão Celular , Humanos , Microscopia Eletrônica , QuebequeRESUMO
A regular progression of polymorphic phase behavior was observed for mixtures of the anionic phospholipid, cardiolipin, and the cationic phospholipid derivative, 1, 2-dioleoyl-sn-glycero-3-ethylphosphocholine. As revealed by freeze-fracture electron microscopy and small-angle x-ray diffraction, whereas the two lipids separately assume only lamellar phases, their mixtures exhibit a symmetrical (depending on charge ratio and not polarity) sequence of nonlamellar phases. The inverted hexagonal phase, H(II,) formed from equimolar mixtures of the two lipids, i.e., at net charge neutrality (charge ratio (CR((+/-))) = 1:1). When one type of lipid was in significant excess (CR((+/-)) = 2:1 or CR((+/-)) = 1:2), a bicontinuous cubic structure was observed. These cubic phases were very similar to those sometimes present in cellular organelles that contain cardiolipin. Increasing the excess of cationic or anionic charge to CR((+/-)) = 4:1 or CR((+/-)) = 1:4 led to the appearance of membrane bilayers with numerous interlamellar contacts, i.e., sponge structures. It is evident that interactions between cationic and anionic moieties can influence the packing of polar heads and hence control polymorphic phase transitions. The facile isothermal, polymorphic interconversion of these lipids may have important biological and technical implications.
Assuntos
Cardiolipinas/química , Lipossomos/química , Fosfolipídeos/química , Animais , Bovinos , Técnica de Fratura por Congelamento , Microscopia Eletrônica , Ácidos Oleicos/química , Fosfatidilcolinas/química , Eletricidade Estática , Relação Estrutura-Atividade , Difração de Raios XRESUMO
Oncostatin M (OSM) is a multifunctional cytokine, a member of the interleukin-6/leukemia inhibitory factor (IL-6/LIF) family, that can regulate a number of connective-tissue cell types in vitro including cartilage and synovial tissue-derived fibroblasts, however its role in joint inflammation in vivo is not clear. We have analyzed murine OSM (muOSM) activity in vitro and in vivo in mouse joint tissue, to determine the potential role of this cytokine in local joint inflammation and pathology. The effects of muOSM and other IL-6/LIF cytokines on mouse synovial fibroblast cultures were assessed in vitro and showed induction of monocyte chemotactic protein-1, interleukin-6, and tissue inhibitor metalloproteinase-1, as well as enhancement of colony growth in soft agarose culture. Other IL-6/LIF cytokines including IL-6, LIF, or cardiotrophin-1, did not have such effects when tested at relatively high concentrations (20 ng/ml). To assess effects of muOSM in articular joints in vivo, we used recombinant adenovirus expressing muOSM cDNA (AdmuOSM) and injected purified recombinant virus (10(6) to 10(8) pfu) intra-articularly into the knees of various mouse strains. Histological analysis revealed dramatic alterations in the synovium but not in synovium of knees treated with the control virus Ad-dl70 or knees treated with Adm-IL-6 encoding biologically active murine IL-6. AdmuOSM effects were characterized by increases in the synovial cell proliferation, infiltration of mononuclear cells, and increases in extracellular matrix deposition that were evident at day 4, but much more marked at days 7, 14, and 21 after administration. The synovium took on characteristics similar to pannus and appeared to contact and invade cartilage. Collectively, these results provide good evidence that OSM regulates synovial fibroblast function differently than other IL-6-type cytokines, and can induce a proliferative invasive phenotype of synovium in vivo in mice on overexpression. We suggest that OSM may contribute to pathology in arthritis.
Assuntos
Fibroblastos/efeitos dos fármacos , Articulação do Joelho/efeitos dos fármacos , Articulação do Joelho/patologia , Peptídeos/farmacologia , Membrana Sinovial/citologia , Sinovite/induzido quimicamente , Adenoviridae/genética , Animais , Divisão Celular/efeitos dos fármacos , Células Cultivadas , Quimiocina CCL2/metabolismo , Fibroblastos/metabolismo , Vetores Genéticos , Interleucina-6/metabolismo , Camundongos , Camundongos Endogâmicos C3H , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos MRL lpr , Oncostatina M , Peptídeos/genética , Proteínas Recombinantes/farmacologiaRESUMO
OBJECTIVE: To determine the ultastructural changes of Taxol (paclitaxel) involution of articular cartilage destruction in collagen induced arthritis (CIA) and to compare with articular cartilage from normal rats. METHODS: Forty-five Louvain rats were randomized to one of 3 protocols for structural analysis: (1) control group, (2) CIA group, and (3) Taxol treated CIA group. The latter group received 10 mg/kg body weight of Taxol at Days 10, 12, and 14 and 7.5 mg/kg body weight of Taxol on Days 16, 18, and 20 postimmunization with collagen type II. Eight days later, each group was examined by light microscopy and scanning and transmission electron microscopy. RESULTS: In Taxol treated rats, the morphology of the articular cartilage reverted to that observed in naive rats except for a striking increase in the thickness of the superficial amorphous layer covering the articular surface. CONCLUSION: The involution of CIA by Taxol suggests that this agent may be useful in the clinical treatment of RA.
Assuntos
Artrite/induzido quimicamente , Artrite/patologia , Cartilagem Articular/efeitos dos fármacos , Cartilagem Articular/ultraestrutura , Colágeno , Paclitaxel/uso terapêutico , Animais , Feminino , Microscopia Eletrônica , Microscopia Eletrônica de Varredura , Ratos , Ratos Endogâmicos , Tarso Animal/efeitos dos fármacos , Tarso Animal/ultraestruturaRESUMO
A novel prism-mirror-prism imaging electron spectrometer with 1 eV energy resolution for a transmission electron microscope permits imaging with spectral energies corresponding to light-optical colour absorptions. The instrument selects the molecular orbital excitations of natural chromophores or of specific dyes normally used in biological light microscopy for delineation and chemical identification, but images them with electron microscopic detail. Heavy atom contrast agents customarily used in electron microscopy are not required. The first results exploit the intrinsic red colour of hematin molecules to demonstrate the potential of the technique and address its spatial resolution. Glycosaminoglycans in cartilage stained with Alcian blue are selectively depicted in situ by means of the electron-induced molecular absorption of this chromophore. Thus, with the use of specific colours the direct or indirect analysis of local chemistry by electron microscopy is possible, and can be carried out with a depiction of spatial detail as small as 16 A, or at least 100-fold finer than observed by light microscopy.
Assuntos
Corantes , Microscopia Eletrônica/métodos , Espectrofotometria/instrumentação , Azul Alciano , Animais , Glicosaminoglicanos/ultraestrutura , Lâmina de Crescimento/ultraestrutura , Hemina/análise , Ratos , Sensibilidade e EspecificidadeRESUMO
Collagen-induced arthritis (CIA) is an animal model of rheumatoid arthritis (RA) that can be regressed with Taxol (paclitaxel), a chemotherapeutic agent. To identify structural changes that occur with involution, the synovium from naive, untreated CIA, and Taxol-treated CIA rats were evaluated by light microscopy plus transmission and scanning electron microscopy. Analysis included detailed images of vascular networks using polymeric corrosion casts. The CIA synovium was morphologically similar to human RA synovium. In CIA, the integrity of the intimal lining is lost by Type-B synoviocytes becoming highly elongated and polarized toward the joint space, resulting in non-overlapping cellular processes and the elimination of the basal lamina. In addition, the lining expanded from a width of 6-10 microns in naives to 200-250 microns in CIA due primarily to increased numbers of both Type-A and -B synoviocytes and more interstitial matrix. Vascular corrosion casts of CIA synovium illustrated a marked increase in blood vessel volume and an extensive interconnecting vascular architecture; neovascular arrays were observed to project toward the synovial surface. In Taxol-treated CIA, the synoviocyte and neovascular components reverted to the naive synovium morphology, suggesting that this agent might be useful in the therapy of RA.
Assuntos
Antineoplásicos Fitogênicos/uso terapêutico , Artrite Reumatoide/tratamento farmacológico , Artrite Reumatoide/patologia , Paclitaxel/uso terapêutico , Animais , Artrite Reumatoide/etiologia , Colágeno/administração & dosagem , Colágeno/imunologia , Molde por Corrosão , Modelos Animais de Doenças , Feminino , Humanos , Microscopia Eletrônica , Microscopia Eletrônica de Varredura , Neovascularização Patológica/tratamento farmacológico , Neovascularização Patológica/etiologia , Neovascularização Patológica/patologia , Ratos , Ratos Endogâmicos , Membrana Sinovial/patologia , Sinovite/tratamento farmacológico , Sinovite/etiologia , Sinovite/patologiaRESUMO
The nasal mucosal is the first site of contact with inhaled antigens. However, the nature of local immune responses and the role of nasal-associated lymphoid tissue (NALT) in those responses have rarely been studied. To characterize the cells involved in mucosally derived immune responses, NALT and Peyer's patch (PP) cells from normal mice, and mice immunized intragastrically or intranasally with cholera toxin (CT), were isolated and analyzed. Compared with PP cells, unstimulated NALT cells contained a higher proportion of T-cells. The CD4:CD8 ratio in NALT cell preparations was less than that observed in PP and more closely resembled that seen in spleen. Additionally, the total B-cell frequency in NALT cell isolates was 20% lower than that observed in PP cell preparations. Although NALT and PP cell isolates contained both mature B-cells and cells undergoing activation to express surface IgA, unlike PP, NALT showed no significant frequency of IgA-switched cells. After intranasal immunization with CT, toxin-specific IgA antibody-forming cells (AFCs) were detected in NALT cell preparations. The numbers of these cells correlated with CT-specific IgA in nasal, but not in gut washes or sera, thus suggesting local nasal production of antigen-specific mucosal antibodies. There was no evidence of anti-CT AFCs in NALT or CT-specific antibody in nasal washes after intragastric CT administration. These results support the notion that nasal mucosal antibody production is best achieved via direct stimulation of IgA-committed, NALT-derived B-cells.
Assuntos
Imunidade nas Mucosas , Mucosa Nasal/imunologia , Nódulos Linfáticos Agregados/imunologia , Alérgenos/imunologia , Animais , Anticorpos Anti-Idiotípicos/análise , Linfócitos B/imunologia , Linfócitos B/ultraestrutura , Relação CD4-CD8 , Toxina da Cólera , Ensaio de Imunoadsorção Enzimática , Feminino , Citometria de Fluxo , Imunoglobulina A/biossíntese , Imunoglobulina A/imunologia , Imunoglobulina G/biossíntese , Imunoglobulina G/imunologia , Tecido Linfoide/imunologia , Tecido Linfoide/ultraestrutura , Camundongos , Camundongos Endogâmicos BALB C , Microscopia Eletrônica de Varredura , Mucosa Nasal/ultraestrutura , Nódulos Linfáticos Agregados/ultraestrutura , Linfócitos T/imunologia , Linfócitos T/ultraestruturaRESUMO
Dentin phosphoproteins are thought to have a primary role in the deposition of mineral on the collagen of dentin. In this study we determined the type of binding between collagen and phosphoproteins necessary for mineral formation onto collagen fibrils and whether the phosphate esters are required. Bovine dentin phosphophoryn or phosvitin from egg yolk were immobilized on reconstituted skin type I collagen fibrils by adsorption or by covalent cross-linking. In some samples the ester phosphate was removed from the covalently cross-linked phosphoproteins by treatment with acid phosphatase. All samples were incubated at 37 degrees C in metastable solutions that do not spontaneously precipitate. Reconstituted collagen fibrils alone did not induce mineral formation. The phosphoproteins adsorbed to the collagen fibrils desorbed when the mineralization medium was added, and mineral was not induced. The mineral induced by the cross-linked phosphoproteins was apatite, and the crystals were confined to the surface of the collagen fibrils. With decreasing medium saturation the time required for mineral induction increased. The interfacial tensions calculated for apatite formation by either phosphoprotein cross-linked to collagen were about the same as that for phosphatidic acid liposomes and hydroxyapatite. This similarity in values indicates that the nucleation potential of these highly phosphorylated surfaces is about the same. It is concluded that phosphoproteins must be irreversibly bound to collagen fibrils for the mineralization of the collagen network in solutions that do not spontaneously precipitate. The phosphate esters of phosphoproteins are required for mineral induction, and the carboxylate groups are not sufficient.
Assuntos
Apatitas/química , Colágeno/química , Dentina/química , Fosfoproteínas/química , Fosfatase Ácida/química , Adsorção , Animais , Sítios de Ligação , Calcificação Fisiológica , Bovinos , Reagentes de Ligações Cruzadas/química , Cristalização , Proteínas do Ovo/química , Microscopia Eletrônica , Fosfoproteínas/ultraestrutura , Fosvitina/química , TemperaturaRESUMO
We have used electron spectroscopic imaging to locate the phosphorus in vaccinia DNA in situ in unstained, ultrathin sections of virions. The phosphorus of the DNA backbone appeared to form a halo on the core periphery surrounding a phosphorus-impoverished central element. These results constrain models for how DNA could be packaged into mature vaccinia particles.
Assuntos
DNA Viral/ultraestrutura , Vaccinia virus/genética , DNA Viral/análise , DNA Viral/química , Microanálise por Sonda Eletrônica , Microscopia Eletrônica , Fósforo/análise , Vaccinia virus/ultraestrutura , Vírion/genética , Vírion/ultraestruturaRESUMO
The spatial and temporal relationships of mineral deposition between matrix vesicles and type I collagen fibrils have been studied in the turkey leg tendon by electron microscopy of cross sections and serial longitudinal thin sections and by electron tomography of longitudinal thick sections. Serial sectioning and electron tomography allow three-dimensional analysis of spatial relationships, overcoming the problems of missing depth information and over-projection of adjacent structures which exist for two-dimensional projections of isolated sections. These techniques reveal that while mineral deposits within matrix vesicles are found remote from calcifying collagen fibrils, the reverse relationship does not occur; all collagen-associated mineral can ultimately be linked to mineral-laden vesicles. These results suggest a temporal sequence of calcification beginning in matrix vesicles and spreading to adjacent collagen fibrils.
Assuntos
Colágeno/fisiologia , Matriz Extracelular/fisiologia , Membro Posterior , Minerais/metabolismo , Tendões/metabolismo , Animais , Processamento de Imagem Assistida por Computador , Microscopia Eletrônica , Tendões/ultraestrutura , Tomografia , PerusRESUMO
'Large' (30-100 microns) and 'small' (10-30 microns) size range taxol-loaded microspheres composed of a blend of biodegradable poly (d,l-lactic acid) (PLA) polymer and nondegradable ethylene-vinyl acetate (EVA) copolymer were prepared using the solvent evaporation method. Encapsulation efficiencies were between 95-100% for taxol in 50:50 EVA:PLA blend microspheres. Between 10-13% of the total taxol content of the microspheres (0.6% w/v taxol loading) was released in 50 days. Using the chick chorioallantoic membrane (CAM) model, the taxol microspheres released sufficient taxol to produce vascular regression and inhibition of angiogenesis. This taxol-loaded microsphere formulation may have potential for the targeted delivery of taxol to a tumor via arterial chemoembolization.
Assuntos
Preparações de Ação Retardada/química , Paclitaxel/administração & dosagem , Animais , Embrião de Galinha , Técnicas In Vitro , Lactatos/química , Ácido Láctico , Microscopia Eletrônica de Varredura , Microesferas , Neovascularização Patológica , Polímeros , Polivinil/químicaRESUMO
The epiphyseal growth plate and articular cartilage matrices were preserved by slam freezing and freeze substitution to optimally retain the native organization for both cellular and matrix components. These specimens were stained and examined using conventional electron microscopic methods. The highly integrated, proteoglycan-rich matrices were examined by computer image analysis using such parameters as distribution, connectivity, orientation, and a variety of morphometric analyses. Also, different aspects of electron tomography and 3D rendering of matrix vesicles and their associated mineral deposits from epiphyseal growth plates and turkey leg tendons are presented.
Assuntos
Cartilagem Articular/ultraestrutura , Matriz Extracelular/ultraestrutura , Lâmina de Crescimento/ultraestrutura , Tendões/ultraestrutura , Animais , Bovinos , Substituição ao Congelamento , Congelamento , Processamento de Imagem Assistida por Computador , Camundongos , Proteoglicanas/ultraestrutura , PerusRESUMO
Electron microscopic studies of calcifying vertebrate tissues reveal the locus of de novo mineral formation within matrix vesicles (MV). The direct involvement of MV in the initiation of mineral formation is supported by the fact that MV isolated from avian growth plate cartilage rapidly accumulate large amounts of Ca2+ and P(i) and induce mineral formation. Exploration of the constituents of MV has revealed two major protein components, a 33 and a 36 kD protein, the former of which binds to cartilage-specific collagens. These annexin-like proteins bind to acidic phospholipids in the presence of submicromolar levels of Ca2+. Antibodies raised against both the purified 33 and the 36 kD MV annexin do not cross-react with the other, indicating that they are distinct proteins. Reported here are studies elucidating the primary structure of both MV proteins using both conventional protein and molecular biologic methods. These studies establish that the 33 kD protein is nearly identical to anchorin CII (annexin V) and that the 36 kD protein is identical to avian annexin II. Immunolocalization studies show that hypertrophic chondrocytes at the calcification front of avian growth plate contain the highest level of these annexins. Further, immunogold labeling indicates that the annexins are localized within MV isolated from the growth plate. Recent studies indicate that annexin V is a new type of ion-selective Ca2+ channel protein that possesses selective collagen binding properties. Since MV are tightly associated with the collagen- and proteoglycan-rich matrix, it is tempting to speculate that this MV protein may be a component of stretch-activated ion channels that enhance Ca2+ uptake during mechanical stress.
Assuntos
Proteínas de Ligação ao Cálcio/química , Lâmina de Crescimento/química , Proteínas de Membrana/química , Proteínas da Gravidez/química , Sequência de Aminoácidos , Animais , Anexina A5 , Anexinas , Proteínas de Ligação ao Cálcio/análise , Proteínas de Ligação ao Cálcio/isolamento & purificação , Galinhas , DNA/química , Eletroforese em Gel de Poliacrilamida , Immunoblotting , Proteínas de Membrana/isolamento & purificação , Microscopia Eletrônica , Dados de Sequência Molecular , Peso Molecular , Proteínas da Gravidez/análise , Proteínas da Gravidez/isolamento & purificaçãoAssuntos
Minerais/metabolismo , Tendões/metabolismo , Animais , Apatitas/metabolismo , Matriz Óssea/metabolismo , Matriz Óssea/ultraestrutura , Cálcio/metabolismo , Colágeno/ultraestrutura , Matriz Extracelular/metabolismo , Matriz Extracelular/ultraestrutura , Modelos Biológicos , Fósforo/metabolismo , Proteoglicanas/metabolismo , Tendões/ultraestrutura , PerusRESUMO
Turkey leg tendons at an early stage of mineralization have been thin sectioned and imaged by electron microscopy. At this stage collagen-associated mineral apatite was found to be present within both the gap and overlap zones. The earliest apatite occurs in a microcrystalline form which gives a rather generalized and characteristic density to both the gap and overlap zones; with subsequent development larger defined apatite crystals arise which span gap/overlap zones. Fourier transformation of such images revealed the major 67 nm axial repeat of the gap/overlap zone plus four other maxima corresponding to repeat spacings of 22, 16, 13, and 11 nm respectively. Computer imaging techniques were used to reconstruct images by using selected spatial frequencies from such transforms. In this manner the subperiodic distributions of mineral were visually enhanced. These subperiodicities are positioned in an asymmetric fashion over the entire D unit repeat aligning with the molecular orientation of the fibril. Analyses of both negatively stained collagen and computer-generated maps of collagen hydrophobicity were compared to the mineral distribution of collagen. Densitometric comparisons showed a positional correlation between the axial banding patterns of mineralized fibrils and those of negatively stained non-mineralized fibrils. Comparable spatial frequencies were also present in transforms between hydrophobic maps and mineral distribution of collagen. These results suggest that the lateral clusterings of hydrophobic residues which span the fibril at specific sites in both the gap and overlap zones serve to prohibit early mineral deposition. This observed hydrophobic influence in combination with the gap space appear as contributing factors in the observed axial distribution of mineral within collagen.
Assuntos
Colágeno/ultraestrutura , Processamento de Imagem Assistida por Computador , Aminoácidos/química , Animais , Densitometria , Perna (Membro) , Minerais/química , Tendões/química , Tendões/ultraestrutura , Perus , Difração de Raios XRESUMO
We have determined the localization of apatite within type I collagen fibrils of calcifying turkey leg tendons by both bright field and selected-area dark field (SADF) electron microscopy and have compared this to computer-modeled, chick type I collagen amino acid sequence data. Apatite crystals occur in both the gap and overlap zones at early stages of mineralization in an asymmetric pattern that corresponds to the polarity, N- to C- orientation, of the collagen molecule. Based on comparisons with computer-generated models of known amino acid sequence of collagen, it was determined for early stages of mineral deposition that apatite is restricted by areas of high hydrophobicity. The gap zone is less hydrophobic than the overlap zone on average but each of these zones had areas of high hydrophobicity that correlated with sites of low localization of mineral. Possible interactions between hydrophobic regions and the process of mineral deposition are discussed.
Assuntos
Apatitas/metabolismo , Colágeno/química , Sequência de Aminoácidos , Animais , Calcificação Fisiológica , Colágeno/ultraestrutura , Simulação por Computador , Microscopia Eletrônica , Tendões/metabolismo , Tendões/ultraestrutura , PerusRESUMO
The morphology of the metaphyseal microvasculature at the epiphysis was examined at both the light and electron microscopic level in rickets and rachitic reversal. The animals studied were normal, rachitic, and rachitic reversed at 8, 24, and 96 hours post-vitamin D administration. The overall architecture of the metaphyseal vessels was significantly altered throughout the intervals examined. In the rachitic animal, arterioles, venules, and capillaries were found adjacent to the growth plate, either directly apposed to the hypertrophic chondrocytes or separated from them by bone-forming cells. These vessels are in many ways similar to the larger arterioles and venules that normally supply the metaphyseal capillary sprouts, but in the normal growing animal are usually located 350-500 microns from the epiphyseal cartilage. The rachitic capillaries appear relatively well differentiated with a partial basement membrane and a perivascular cell lining. In early rachitic reversal, small vascular projections are induced to grow from the large diameter venules that border upon the hypertrophic chondrocytes. These vascular sprouts that invade the epiphyseal cartilage are quite undifferentiated, with no basement membrane or pericyte lining at the sprout apex and occasional abluminal endothelial cell projections. Within 96 hours, the metaphyseal microvasculature has returned to an apparently normal state with only capillaries at the cartilage-vascular interface and larger vessels (arterioles and venules) located several hundred microns deeper into the metaphysis. The sequential processes of differentiation and cessation of capillary growth followed by dedifferentiation and reinitiation of microvascular growth make the rachitic system a unique one in which to study angiogenesis.
Assuntos
Lâmina de Crescimento/irrigação sanguínea , Neovascularização Patológica/patologia , Raquitismo/patologia , Animais , Arteríolas/efeitos dos fármacos , Arteríolas/ultraestrutura , Membrana Basal/efeitos dos fármacos , Membrana Basal/ultraestrutura , Capilares/efeitos dos fármacos , Capilares/ultraestrutura , Modelos Animais de Doenças , Lâmina de Crescimento/efeitos dos fármacos , Lâmina de Crescimento/patologia , Injeções Subcutâneas , Masculino , Microcirculação , Microscopia Eletrônica , Ratos , Fatores de Tempo , Vênulas/efeitos dos fármacos , Vênulas/ultraestrutura , Vitamina D/administração & dosagem , Vitamina D/farmacologiaRESUMO
This study is concerned with the nature and distribution of mineral in the gallbladder of a patient with chronic cholecystitis. Light and electron microscopic imaging revealed the mineral to be in the epithelial cells of the mucosa and fibroblasts of the submucosa. In the epithelial cells at the early stages of deposition, mineral was located in the nuclei and throughout the cytoplasm in association with interdigitating cell processes and apical microvilli but was absent in mitochondria. Elemental and electron diffraction analyses indicated the mineral inclusions to be apatite in nature.
Assuntos
Apatitas/análise , Calcinose/metabolismo , Doenças da Vesícula Biliar/metabolismo , Adulto , Calcinose/complicações , Calcinose/patologia , Núcleo Celular/química , Colecistite/complicações , Citoplasma/química , Epitélio/química , Feminino , Fibroblastos/química , Vesícula Biliar/química , Vesícula Biliar/patologia , Doenças da Vesícula Biliar/patologia , Humanos , Microscopia Eletrônica , Mucosa/química , Distribuição TecidualRESUMO
This study is concerned with the cryogenic preservation of intrafibrillar apatite distribution in type I collagen of turkey leg tendons. Cryogenic specimen preparations by the rapid freezing of nonfixed and noncryoprotected leg tendons were performed by two different protocols: (1) low temperature substitution, fixation and staining followed by low temperature embedment; (2) frozen hydrated and air-dried cryosections were examined with the electron microscope at -165 degrees C and normal operating temperatures, respectively. These protocols revealed the axial periodicity for mineralized collagen to have a 65-69 nm range with a mean value of 67 nm as determined by point-to-point measurements. Mineral distributions and specific apatite visualization were examined by electron microscopic imaging in bright field and selected-area dark field, respectively. Fourier filtered images and image subtraction were used to separate the axial repeating and nonrepeating intrafibrillar mineral domains of collagen. The removal of these axial repeats revealed an underlying and integrated mineral distribution, demonstrating that apatite is not confined to axial periodicities such as those of the gap zone.
Assuntos
Colágeno/metabolismo , Minerais/metabolismo , Tendões/metabolismo , Animais , Apatitas/metabolismo , Colágeno/análise , Colágeno/ultraestrutura , Criopreservação , Processamento de Imagem Assistida por Computador , Microscopia Eletrônica/métodos , Minerais/análise , Tendões/química , Tendões/ultraestrutura , PerusRESUMO
Turkey leg tendons were used as a model tissue to study the spatial and temporal relationships of mineral deposition between matrix vesicles and collagen fibrils by various electron microscopic techniques--bright field, selected-area dark field (SADF), and electron spectroscopic imaging (ESI). These latter imaging techniques enabled the direct localization and spatial distributions of both apatite crystals and atomic elements (Ca, P) within matrix vesicles and collagen. In longitudinal planes of section, a consistent vectorial gradient of mineralization was observed which started with the first localization of apatite mineral in matrix vesicles; with further development, the mineral spread from the vesicle to the extravesicular interstices and then into the adjacent collagen fibrils. Once intrafibrillar, the mineral was observed to advance both laterally and axially. The association of vesicle/collagen mineral was examined by ESI analysis of Ca and P elemental maps and appeared as a continuum between the vesicles and the adjacent collagen fibrils. Similarly, an intimate spatial relationship was observed between the mineral of vesicles and collagen in transversely cut sections of tendon. The sequential development of this mineralized matrix is discussed in light of matrix vesicle/collagen interactions.