RESUMO
Collagenous tissues exhibit anisotropic optical properties such as birefringence and linear dichroism (LD) as a result of their structurally oriented supraorganization from the nanometer level to the collagen bundle scale. Changes in macromolecular order and in aggregational states can be evaluated in tendon collagen bundles using polarization microscopy. Because there are no reports on the status of the macromolecular organization in tendon explants, the objective of this work was to evaluate the birefringence and LD characteristics of collagen bundles in rat calcaneal tendons cultivated in vitro on substrates that differ in their mechanical stiffness (plastic vs. glass) while accompanying the expected occurrence of cell migration from these structures. Tendon explants from adult male Wistar rats were cultivated for 8 and 12 days on borosilicate glass coverslips (n = 3) and on nonpyrogenic polystyrene plastic dishes (n = 4) and were compared with tendons not cultivated in vitro (n = 3). Birefringence was investigated in unstained tendon sections using high-performance polarization microscopy and image analysis. LD was studied under polarized light in tendon sections stained with the dichroic dyes Ponceau SS and toluidine blue at pH 4.0 to evaluate the orientation of proteins and acid glycosaminoglycans (GAG) macromolecules, respectively. Structural remodeling characterized by the reduction in the macromolecular orientation, aggregation and alignment of collagen bundles, based on decreased average gray values concerned with birefringence intensity, LD and morphological changes, was detected especially in the tendon explants cultivated on the plastic substrate. These changes may have facilitated cell migration from the lateral regions of the explants to the substrates, an event that was observed earlier and more intensely upon tissue cultivation on the plastic substrate. The axial alignment of the migrating cells relative to the explant, which occurred with increased cultivation times, may be due to the mechanosensitive nature of the tenocytes. Collagen fibers possibly played a role as a signal source to cells, a hypothesis that requires further investigation, including studies on the dynamics of cell membrane receptors and cytoskeletal organization, and collagen shearing electrical properties.
Assuntos
Tendão do Calcâneo , Ratos , Masculino , Animais , Ratos Wistar , Microscopia de Polarização , Colágeno/metabolismo , PlásticosRESUMO
Toluidine blue (TB) staining either alone or in association with other methodologies has the potential to answer a variety of biological questions regarding the human, animal and plant tissues or cells. In this brief review, we not only report the primary use of TB to detect the anionic substrates and availability of their binding sites, but also unveil the resulting applications of TB staining in biological research. Among these applications, the uses of TB staining to identify the changes in chromatin DNA-protein complexes, nucleolus location, and extracellular matrix proteoglycan complexes associated with different physiological and pathological events are described. The usefulness of TB staining to monitor the effects elicited by environmental insults on chromatin and intercalation of drugs into the DNA is also included.
Assuntos
Cromatina/metabolismo , DNA , Coloração e Rotulagem , Cloreto de Tolônio , Animais , Sítios de Ligação/fisiologia , Humanos , Proteoglicanas/metabolismo , Coloração e Rotulagem/métodosRESUMO
The larvae of the two distantly related nonsocial bees Ericrocis lata (Apidae) and Hesperapis (Carinapis) rhodocerata (Melittidae), which develop mostly under arid desert areas of North America, and that differ in that they either spin (E. lata) or do not spin (H. rhodocerata) protective cocoons before entering diapause, produce transparent films that cover the larval integument. To understand the nature of these films, their responses to topochemical tests and their characteristics when examined with fluorescence and high-performance polarization microscopy and microspectroscopy were studied. A positive staining by Sudan black B, birefringence of negative sign, and a Fourier transform-infrared (FT-IR) spectrum typical of lipids were detected for the integument covering of both species. The FT-IR signature, particularly, suggests a wax chemical composition for these lipid coverings, resembling the waxes that are used as construction materials in the honey cells produced by social bees. Considering the arid environmental conditions under which these larvae develop, we hypothesize that their covering films may have evolved as protection against water depletion. This hypothesis seems especially appropriate for H. rhodocerata larvae, which are capable of undergoing a long diapause period in the absence of a protective cocoon.
Assuntos
Abelhas/fisiologia , Diapausa/fisiologia , Tegumento Comum/fisiologia , Microscopia de Polarização/métodos , Espectroscopia de Infravermelho com Transformada de Fourier/métodos , AnimaisRESUMO
Valproic acid (VPA), a well-known histone deacetylase inhibitor, has been reported to affect the DNA methylation status in addition to inducing histone hyperacetylation in several cell types. In HeLa cells, VPA promotes histone acetylation and chromatin remodeling. However, DNA demethylation was not checked in this cell model for standing effects longer than those provided by histone acetylation, which is a rapid and transient phenomenon. Demonstration of VPA-induced DNA demethylation in HeLa cells would contribute to understanding the effect of VPA on an aggressive tumor cell line. In the present work, DNA demethylation in VPA-treated HeLa cells was assessed by image analysis of chromatin texture, the abundance of 5-methylcytosine (5mC) immunofluorescence signals and Fourier transform-infrared (FT-IR) microspectroscopy centered on spectral regions related to the vibration of-CH3 groups. Image analysis indicated that increased chromatin unpacking promoted by a 4-h-treatment with 1.0 mM VPA persisted for 24 h in the absence of the drug, suggesting the occurrence of DNA demethylation that was confirmed by decreased 5mC immunofluorescence signals. FT-IR spectra of DNA samples from 1 mM or 20 mM VPA-treated cells subjected to a peak fitting analysis of the spectral window for-CH3 stretching vibrations showed decreased vibrations and energy of these groups as a function of the decreased abundance of 5mC induced by increased VPA concentrations. Only the 20 mM-VPA treatment caused an increase in the ratio of -CH3 bending vibrations evaluated at 1375 cm-1 in relation to in-plane vibrations of overall cytosines evaluated at 1492 cm-1. CH3 stretching vibrations showed to be more sensitive than-CH3 bending vibrations, as detected with FT-IR microspectroscopy, for studies aiming to associate vibrational spectroscopy and changes in DNA 5mC abundance.
Assuntos
Metilação de DNA , Ácido Valproico/farmacologia , Células HeLa , Humanos , Microscopia de Fluorescência , Espectroscopia de Infravermelho com Transformada de Fourier , Análise Espectral/métodos , VibraçãoRESUMO
Rat ear cartilage was studied using Fourier transform-infrared (FT-IR) microspectroscopy to expand the current knowledge which has been established for relatively more complex cartilage types. Comparison of the FT-IR spectra of the ear cartilage extracellular matrix (ECM) with published data on articular cartilage, collagen II and 4-chondroitin-sulfate standards, as well as of collagen type I-containing dermal collagen bundles (CBs) with collagen type II, was performed. Ear cartilage ECM glycosaminoglycans (GAGs) were revealed histochemically and as a reduction in ECM FT-IR spectral band heights (1140-820 cm-1) after testicular hyaluronidase digestion. Although ear cartilage is less complex than articular cartilage, it contains ECM components with a macromolecular orientation as revealed using polarization microscopy. Collagen type II and GAGs, which play a structural role in the stereo-arrangement of the ear cartilage, contribute to its FT-IR spectrum. Similar to articular cartilage, ear cartilage showed that proteoglycans add a contribution to the collagen amide I spectral region, a finding that does not recommend this region for collagen type II quantification purposes. In contrast to articular cartilage, the symmetric stretching vibration of -SO3- groups at 1064 cm-1 appeared under-represented in the FT-IR spectral profile of ear cartilage. Because the band corresponding to the asymmetric stretching vibration of -SO3- groups (1236-1225 cm-1) overlapped with that of amide III bands, it is not recommended for evaluation of the -SO3- contribution to the FT-IR spectrum of the ear cartilage ECM. Instead, a peak (or shoulder) at 1027-1016 cm-1 could be better considered for this intent. Amide I/amide II ratios as calculated here and data from the literature suggest that protein complexes of the ear cartilage ECM are arranged with a lower helical conformation compared to pure collagen II. The present results could motivate further studies on this tissue under pathological or experimental states involving ear cartilage.
Assuntos
Cartilagem Articular/metabolismo , Cartilagem da Orelha/metabolismo , Matriz Extracelular/metabolismo , Espectroscopia de Infravermelho com Transformada de Fourier , Animais , Sulfatos de Condroitina/metabolismo , Colágeno Tipo I/metabolismo , Colágeno Tipo II/metabolismo , Glicosaminoglicanos/metabolismo , RatosRESUMO
Entheses are specialized biological structures that functionally anchor tendons to bones. The complexity, mechanical characteristics and properties of the entheses, particularly those related to exercise, mechanical load and pathologies, have been extensively analyzed; however, the macromolecular organization of the enthesis fibers, as assessed by polarization microscopy, has not yet been investigated. Morphological and optical anisotropy characteristics, such as birefringence, linear dichroism (LD) and differential interference contrast (DIC-PLM) properties, are thus analyzed in this study of a healthy adult mouse calcaneal tendon-bone enthesis. The molecular and supramolecular order of collagen and GAGs was determined for the collagen bundles of this enthesis. Based on a birefringence plot pattern as well as on metachromasy and linear dichroism after toluidine blue staining at pH 4.0, a similarity between the calcaneal tendon-bone enthesis and cartilage during ossification may be assumed. This similarity is assumed to favor the adequacy of this enthesis to support a compressive load. Considering that the collagen-proteoglycan complexes and the enthesis fibers themselves have a chiral nature, these structures could be acting via reciprocal signaling with the cellular environment of the enthesis.
Assuntos
Anisotropia , Mediadores da Inflamação/imunologia , Fenômenos Ópticos , Animais , Camundongos , Engenharia TecidualRESUMO
Although several treatments for tendon lesions have been proposed, successful tendon repair remains a great challenge for orthopedics, especially considering the high incidence of re-rupture of injured tendons. Our aim was to evaluate the pharmacological potential of Aloe vera on the content and arrangement of glycosaminoglycans (GAGs) during tendon healing, which was based on the effectiveness of A. vera on collagen organization previously observed by our group. In rats, a partial calcaneal tendon transection was performed with subsequent topical A. vera application at the injury site. The tendons were treated with A. vera ointment for 7 days and excised on the 7(th) , 14(th) , or 21(st) day post-surgery. Control rats received ointment without A. vera. A higher content of GAGs and a lower amount of dermatan sulfate were detected in the A. vera-treated group on the 14(th) day compared with the control. Also at 14 days post-surgery, a lower dichroic ratio in toluidine blue stained sections was observed in A. vera-treated tendons compared with the control. No differences were observed in the chondroitin-6-sulfate and TGF-ß1 levels between the groups, and higher amount of non-collagenous proteins was detected in the A. vera-treated group on the 21(st) day, compared with the control group. No differences were observed in the number of fibroblasts, inflammatory cells and blood vessels between the groups. The application of A. vera during tendon healing modified the arrangement of GAGs and increased the content of GAGs and non-collagenous proteins.
Assuntos
Aloe , Glicosaminoglicanos/metabolismo , Fitoterapia , Preparações de Plantas/uso terapêutico , Traumatismos dos Tendões/tratamento farmacológico , Tendões/efeitos dos fármacos , Cicatrização/efeitos dos fármacos , Administração Tópica , Animais , Ensaio de Imunoadsorção Enzimática , Masculino , Preparações de Plantas/administração & dosagem , Ratos , Ratos Wistar , Traumatismos dos Tendões/metabolismo , Tendões/metabolismoRESUMO
BACKGROUND: Lower levels of cytosine methylation have been found in the liver cell DNA from non-obese diabetic (NOD) mice under hyperglycemic conditions. Because the Fourier transform-infrared (FT-IR) profiles of dry DNA samples are differently affected by DNA base composition, single-stranded form and histone binding, it is expected that the methylation status in the DNA could also affect its FT-IR profile. METHODOLOGY/PRINCIPAL FINDINGS: The DNA FT-IR signatures obtained from the liver cell nuclei of hyperglycemic and normoglycemic NOD mice of the same age were compared. Dried DNA samples were examined in an IR microspectroscope equipped with an all-reflecting objective (ARO) and adequate software. CONCLUSIONS/SIGNIFICANCE: Changes in DNA cytosine methylation levels induced by hyperglycemia in mouse liver cells produced changes in the respective DNA FT-IR profiles, revealing modifications to the vibrational intensities and frequencies of several chemical markers, including νas -CH3 stretching vibrations in the 5-methylcytosine methyl group. A smaller band area reflecting lower energy absorbed in the DNA was found in the hyperglycemic mice and assumed to be related to the lower levels of -CH3 groups. Other spectral differences were found at 1700-1500 cm(-1) and in the fingerprint region, and a slight change in the DNA conformation at the lower DNA methylation levels was suggested for the hyperglycemic mice. The changes that affect cytosine methylation levels certainly affect the DNA-protein interactions and, consequently, gene expression in liver cells from the hyperglycemic NOD mice.
Assuntos
Metilação de DNA , DNA/química , DNA/metabolismo , Diabetes Mellitus Experimental/metabolismo , Fígado/patologia , Espectroscopia de Infravermelho com Transformada de Fourier , Animais , Citosina/metabolismo , Diabetes Mellitus Experimental/genética , Diabetes Mellitus Experimental/patologia , Feminino , Camundongos , Camundongos Endogâmicos NODRESUMO
Analysis of the formation of extended chromatin fibers (ECFs) in response to the action of gravity following lysis by hypertonic and detergent solutions is a useful technical procedure relevant for studies of the positioning of particular DNA signals on chromatin filaments. Additionally, if toluidine blue molecules are allowed to bind electrostatically to available DNA phosphates on ECFs, the birefringence brightness generated in these filaments, as observed by polarization microscopy, facilitates the description of the frequency of ECF formation and extension of the chromatin filaments generated. Thus, different patterns of DNA-nuclear matrix protein associations related to varying transcriptional activities and chromatin organization in isolated cells can be assessed. A technique for producing ECFs in different isolated cell types under variable physiological and/or pathological conditions is detailed in this chapter.
Assuntos
Cromatina/metabolismo , Microscopia de Polarização/métodos , Animais , Birrefringência , Hepatócitos/citologia , Camundongos , Coloração e RotulagemRESUMO
Chromatin packaging plays a significant role in regulating gene transcription. Study of the higher-order packing states of chromatin by image analysis at the light microscope level, especially when validated by methods of molecular biology, immunochemistry, and/or immunocytochemistry, enabled the detection of changes involved in the processes associated with or preceding alterations in transcriptional activities. Here, we recommend and describe the use of relatively simple methods for staining and detecting chromatin remodelling by image analysis.
Assuntos
Montagem e Desmontagem da Cromatina , Processamento de Imagem Assistida por Computador/métodos , Animais , Cromatina , Camundongos , Microespectrofotometria , Células NIH 3T3 , Fenótipo , Coloração e RotulagemRESUMO
The Malpighian tubule cell nuclei of male Panstrongylus megistus, a vector of Chagas disease, contain one chromocenter, which is composed solely of the Y chromosome. Considering that different chromosomes contribute to the composition of chromocenters in different triatomini species, the aim of this study was to determine the contribution of AT-, GC-, and methylated cytidine-rich DNA in the chromocenter as well as in euchromatin of Malpighian tubule cell nuclei of P. megistus in comparison with published data for Triatoma infestans. Staining with 4',6-diamidino-2-phenylindole/actinomycin D and chromomycin A(3)/distamycin, immunodetection of 5-methylcytidine and AgNOR test were used. The results revealed AT-rich/GC-poor DNA in the male chromocenter, but equally distributed AT and GC DNA sequences in male and female euchromatin, like in T. infestans. Accumulation of argyrophilic proteins encircling the chromocenter did not always correlate with that of GC-rich DNA. Methylated DNA identified by immunodetection was found sparsely distributed in the euchromatin of both sexes and at some points around the chromocenter edge, but it could not be considered responsible for chromatin condensation in the chromocenter, like in T. infestans. However, unlike in T. infestans, no correlation between the chromocenter AT-rich DNA and nucleolus organizing region (NOR) DNA was found in P. megistus.
Assuntos
Sequência Rica em At , Cromatina/genética , Metilação de DNA , Sequência Rica em GC , Túbulos de Malpighi/citologia , Panstrongylus/genética , Animais , Núcleo Celular/genética , Células Cultivadas , Citidina/análogos & derivados , Citidina/metabolismo , Células Epiteliais/metabolismo , Feminino , Genes de Insetos , Masculino , Camundongos , Região Organizadora do Nucléolo/metabolismo , Ninfa/citologia , Ninfa/genética , Panstrongylus/citologiaRESUMO
Tendons are formed by dense connective tissue composed of an abundant extracellular matrix (ECM) that is constituted mainly of collagen molecules, which are organized into fibrils, fibers, fiber bundles and fascicles helicoidally arranged along the largest axis of the tendon. The biomechanical properties of tendons are directly related to the organization of the collagen molecules that aggregate to become a super-twisted cord. In addition to collagen, the ECM of tendons is composed of non-fibrillar components, such as proteoglycans and non-collagenous glycoproteins. The capacity of tendons to resist mechanical stress is directly related to the structural organization of the ECM. Collagen is a biopolymer and presents optical anisotropies, such as birefringence and linear dichroism, that are important optical properties in the characterization of the supramolecular organization of the fibers. The objective of this study was to present a review of the composition and organization of the ECM of tendons and to highlight the importance of the anisotropic optical properties in the study of alterations in the ECM.
Assuntos
Matriz Extracelular/química , Matriz Extracelular/ultraestrutura , Tendões/química , Tendões/ultraestrutura , Animais , Anisotropia , Humanos , Fenômenos MecânicosRESUMO
Few studies have analyzed the effect of stretching after immobilization on the structural and biochemical properties of tendons. Here, the effect of stretching and immobilization on the proximal (p), intermediate (i), and distal (d) regions of the deep digital flexor tendon in rats was analyzed. The d region was subjected to compression and tension forces, the i region was subjected to compressive forces and the p region received tension forces. Rats were separated into five groups: GI--control for GII; GII--immobilized rats; GIII--control for GIV and GV groups; GIV--immobilized and stretched rats; and GV--immobilized rats which were allowed free cage activity. GII showed a higher molecular organization in the d and p regions as detected by measuring optical retardation, a lower concentration of hydroxyproline in the i region and a significant decrease in noncollagenous proteins found in the three regions of the tendon. Regarding the glycosaminoglycans, diminishing dermatan sulfate and the absence of chondroitin sulfate in the i region were observed in GII when compared to GI. However, in the same region of GIV, higher concentrations of chondroitin and dermatan sulfate were observed along with a strong metachromasy. An increase in hydroxyproline content in the i region and a higher molecular organization in the d and p regions were observed in GIV. Apparently, the active isoforms of metalloproteinase-2 also increased after stretching in all regions. These results suggest that stretching after immobilization contributed to the increase in molecular organization and to the synthesis of extracellular matrix components.
Assuntos
Imobilização , Exercícios de Alongamento Muscular , Tendões/fisiologia , Animais , Eletroforese em Gel de Ágar , Eletroforese em Gel de Poliacrilamida , Colágenos Fibrilares/metabolismo , Hidroxiprolina/metabolismo , Masculino , Aparelhos Ortopédicos , Proteólise , Ratos , Ratos Wistar , Extratos de TecidosRESUMO
Wings of the butterflies Morpho aega and Eryphanis reevesi were investigated in the present study by fluorescence, polarization and infra-red (IR) spectroscopic microscopy with the aim of identifying the oriented organization of their components and morphological details of their substructures. These wings were found to exhibit a strong iridescent glow depending on the angle of the incident light; their isolated scales exhibited blue fluorescence. Parallel columns or ridges extend from the pad and sockets to the dented apical scale's region, and they are perpendicular to the ribs that connect the columnar ridges. The scales reveal linear dichroism (LD) visually, when attached on the wing matrix or isolated on slides. The LD was inferred to be textural and positive and was also demonstrated with IR microscopy. The scale columns and ribs are birefringent structures. Images obtained before and after birefringence compensation allowed a detailed study of the scale morphology. Form and intrinsic birefringence findings here estimated and discussed in the context of nonlinear optical properties, bring to the level of morphology the state of molecular order and periodicity of the wing structure. FT-IR absorption peaks were found at wavenumbers which correspond to symmetric and asymmetric (-N-H) stretching, symmetric (-C-H) stretching, amide I (-CO) stretching, amide II(-N-H), and ß-linking. Based on LD results obtained with polarized IR the molecular vibrations of the wing scales of M. aega and E. reevesi are assumed to be oriented with respect to the long axis of these structures.
Assuntos
Birrefringência , Borboletas/ultraestrutura , Microscopia , Fenômenos Ópticos , Asas de Animais/ultraestrutura , AnimaisRESUMO
Lysyl oxidases (LOXs) are enzymes that permit the covalent crosslinking of the component chains of collagen and elastin. These enzymes are present inside the nuclei of certain mammalian cells. Previous studies have proposed LOX binding to histone H1 in vitro, and histone H1 is known to control global chromatin compaction and mitotic chromosome architecture. Therefore, in the present study, we analyzed chromatin supraorganizational changes, mitotic abnormalities, mitotic indices and cell death ratios in COS-7 and NRK-49F cells with high and low lox expression levels, respectively. The objective was to support biochemical data of LOX-H1 interaction, by providing evidence of chromatin remodeling in vivo, under different lox expressions. Chromatin decondensation assessed by image analysis was observed in COS-7 cells with increased lox expression. This decondensation is suggested to be promoted by LOX actions on histone H1, which loosens the DNA-H1 complex. In NRK-49F cells transfected with antisense lox or subjected to treatment with beta-aminopropionitrile (BAPN), chromatin condensation and nuclear phenotypic variability were found, which may be due to reduced LOX-H1 interaction. When lox expression was increased in COS-7 cells, the frequency of irregular chromosome plates was not affected, but cell proliferation decreased and "cell death preceded by multinucleation" increased. In NRK-49F cells there was accelerated proliferation induced by transfection with the antisense lox, and confirmed when cells were treated with BAPN. Apoptosis increased in NRK-49F cells only with BAPN treatment whereas cell death preceded by multinucleation increased only after antisense lox transfection. The data presented herein regarding chromatin remodeling indirectly support the hypothesis that LOX binds to histone H1 in vivo. Cell proliferation in COS-7 and NRK-49F cells and cell death at least in COS-7 cells agree with predicted effects of LOX interference in these processes.
Assuntos
Cromatina/metabolismo , Histonas/metabolismo , Mitose , Mapeamento de Interação de Proteínas , Proteína-Lisina 6-Oxidase/metabolismo , Animais , Morte Celular , Linhagem Celular , Proliferação de Células , Chlorocebus aethiops , Montagem e Desmontagem da Cromatina , Expressão Gênica , RatosRESUMO
Collagen fiber structure and organization have been found to vary in different tendon types. Differences have been reported in the FT-IR spectra of the amide I band of collagen-containing structures. In the present study, the FT-IR spectral characteristics of the amide I band of the bovine flexor tendon and the extended rat tail tendon were compared by using the diamond attenuated total reflectance technique. The objective was to associate FT-IR spectral characteristics in tendons with their different collagen fiber supraorganization and biomechanical properties. Nylon 6 and poly-L-lysine were used as polyamide models. Each of these materials was found to exhibit molecular order and crystallinity, as revealed by their birefringence. The following FT-IR parameters were evaluated: amide I band profile, absorption peaks and areas, and the 1655 cm⻹/1690 cm⻹ absorbance ratio. The amide I area and the 1655 cm⻹/1690 cm⻹ absorbance ratio were significantly higher for the bovine flexor tendon, indicating that its collagen fibers are richer in pyridinoline-type cross-linking, proline and/or hydroxyproline and H-bonding, and that these fibers are more packed and supraorganizationally ordered than those in the rat tail tendon. This conclusion is additionally supported by differences in collagen solubility and biochemical/biomechanical properties of the tendons.
Assuntos
Amidas/química , Colágeno Tipo I/química , Espectroscopia de Infravermelho com Transformada de Fourier , Animais , Anisotropia , Bovinos , Microscopia de Polarização , RatosRESUMO
Optical anisotropic characteristics investigated by polarization microscopy have been valuable for the study of the oriented organization of collagen fibers in tendons. However, topographic differences in supramolecular organization of collagen fibers along extensive areas in tendons have not yet been described. Here, the statistical variability of the oriented organization of collagen fibers along extensive areas (10(5)-10(6)microm(2)) of 7-microm thick unstained sections of rat calcaneal tendons were studied by assessing their birefringence with polarization microscopy and image analysis, and the periodic frequency distribution in their birefringent images with the fast Fourier transform (FFT). Various levels of birefringence intensity were determined principally by image analysis procedures, and periodicity in the birefringent images was revealed by FFT line profile and spectrum image patterns. Present results support the idea of a helical distribution for collagen bundles along the tendon long axis, and of a statistical architecture for the rat calcaneal tendons in terms of variability in the oriented distribution of their collagen fibers.
Assuntos
Anisotropia , Colágeno/metabolismo , Tendões/anatomia & histologia , Tendões/metabolismo , Animais , Birrefringência , Análise de Fourier , Masculino , Microscopia de Polarização , Ratos , Ratos WistarRESUMO
The structural and supraorganizational arrangement of the chordae tendineae components is very important for a better understanding of their morphophysiological relationships. This study aims to evaluate the degree of statistical variability of the distribution and orientation of collagen fibers and their undulations (crimps), in porcine chordae tendineae. Polarization microscopy, in association with image analysis, was used for the analysis of birefringent images and detection of surface plots, Fast Fourier transforms, and form birefringence curve profiles. A marked variability in the collagen fiber's twisted and intertwined orientation was found not only along the long axis of the chordae tendineae but also in their 3-D structure, including that of the crimp structures. Crimp was demonstrated to not represent a homogeneous distribution of bands; the best methods to quantify its variability were the Fast Fourier transform and the line profile extended along the long axis of the chordae tendineae. Collagen fiber rings, considered to possibly protecting the integrity of the chordae tendineae, were observed to wrap around them. A statistically helical structure is assumed for the supraorganization of the collagen fibers in the chordae tendineae, which is suggested here to be a chiral body.
Assuntos
Cordas Tendinosas/química , Colágeno/química , Processamento de Imagem Assistida por Computador/métodos , Microscopia de Polarização/métodos , Animais , Anisotropia , Análise de Fourier , Conformação Proteica , SuínosRESUMO
Chromatin supraorganization and extensibility and nuclear glycoprotein content have been reported to change in hepatocytes from mice during development and aging, as well as under starvation and refeeding conditions. In non-obese diabetic (NOD) mice, the expression of insulin-dependent diabetes may be accompanied by metabolic changes in the liver. These changes are likely to be similar to those involved in the aging processes of non-diabetic animals. Therefore, we hypothesized that the chromatin organization, as well as the physical properties and compositions of hepatocyte nuclei would also be affected in NOD mice in the same way as those in aged non-diabetic mice. Nuclear image parameters were evaluated by image analysis of Feulgen-stained preparations. Chromatin extensibility in response to gravity was observed with polarized light after lysis and toluidine blue staining. The Con-A response of nuclear glycoproteins was evaluated with scanning microspectrophotometry. These characteristics were assessed using hepatocyte imprints from female NOD mice after a 28-day period of diabetes expression. Observations and measurements were made in comparison to healthy BALB/c mice. Total RNA amounts were determined for livers of NOD and BALB/c mice. Enhanced polyploidy levels, a decrease in chromatin higher-order packing states, an increased frequency of extended chromatin fiber formation, and deeper Con-A-responsive chromatin areas were observed in the hepatocytes of the NOD mice expressing insulin-dependent diabetes. Reduced amounts of total RNA were also found in the livers of these mice. Our findings for NOD mice expressing insulin-dependent diabetes are consistent with previously reported data for old-aged mice of the inbred strain A/Uni and may reflect changes in transcriptional activities associated with the stressful physiological demands on the liver during the expression of diabetes.
Assuntos
Núcleo Celular/química , Núcleo Celular/ultraestrutura , Cromatina/ultraestrutura , DNA/análise , Glicoproteínas/análise , Hepatócitos/química , Hepatócitos/ultraestrutura , RNA/análise , Animais , Diabetes Mellitus/patologia , Fígado/patologia , Camundongos , Camundongos Endogâmicos BALB CRESUMO
Os avanços recentes da Odontologia, aliados ao aprimoramento dos materiais obtidos através das técnicas de bioengenharia, impulsionaram o desenvolvimento de um novo substituto ósseo, composto por colágeno bovino tipo I e hidroxiapatita. Os autores acompanharam 15 pacientes com diferentes diagnósticos e nos quais foi utilizado o material em questão. Os resultados favoráveis encontrados estiveram de acordo com os resultados de estudo histopatológico e foram atribuídos às características de biocompatibilidade, osteocondução e osteoindução e correlacionados com dados da literatura científica.