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Effects of different blood components on clot retraction analysed by measuring elasticity with a free oscillating rheometer.

Free oscillation rheometry (FOR) using the ReoRox 4 instrument makes it possible, at bedside, to study the coagulation process in blood over time and gives information on clotting time and coagulum elastic properties. In order to find out how various factors influence the FOR analysis we studied the coagulation process and change of elasticity over time in non-anticoagulated and citrated blood samples, plasma samples with various platelet concentrations (0-200 x 109/l) and blood samples with various haematocrit (0-40%). Blood samples supplemented with fibrinogen were analysed to elucidate the importance of fibrinogen on elasticity. The importance of the GPIIb/IIIa receptor on platelets was investigated by comparing the elasticity development in blood samples in presence and absence of a GPIIb/IIIa receptor inhibitor, abciximab. Anticoagulation with citrate did not have major influence on the viscoelastic properties of the coagulum. Increasing number of platelets and increasing fibrinogen concentration resulted in higher elasticity while increasing haematocrit gave lower elasticity. Blood samples with GPIIb/IIIa receptor inhibitor had very low elasticity indicating the importance of functional GPIIb/IIIa receptors. In conclusion we consider FOR to be a useful method to study the elastic properties of the coagulum. Various factors such as the number of red blood cells and platelets as well as the fibrinogen concentration should be taken into consideration when evaluating the results. The ReoRox 4 instrument had excellent measuring range and unusually small artefactual effects on clot elasticity induced by the instrument in comparison with published results on other instruments.

Blood Platelets↗

Elastic filaments in situ in cardiac muscle: deep-etch replica analysis in combination with selective removal of actin and myosin filaments.

To clarify the full picture of the connectin (titin) filament network in situ, we selectively removed actin and myosin filaments from cardiac muscle fibers by gelsolin and potassium acetate treatment, respectively, and observed the residual elastic filament network by deep-etch replica electron microscopy. In the A bands, elastic filaments of uniform diameter (6-7 nm) projecting from the M line ran parallel, and extended into the I bands. At the junction line in the I bands, which may correspond to the N2 line in skeletal muscle, individual elastic filaments branched into two or more thinner strands, which repeatedly joined and branched to reach the Z line. Considering that cardiac muscle lacks nebulin, it is very likely that these elastic filaments were composed predominantly of connectin molecules; indeed, anti-connectin monoclonal antibody specifically stained these elastic filaments. Further, striations of approximately 4 nm, characteristic of isolated connectin molecules, were also observed in the elastic filaments. Taking recent analyses of the structure of isolated connectin molecules into consideration, we concluded that individual connectin molecules stretched between the M and Z lines and that each elastic filament consisted of laterally-associated connectin molecules. Close comparison of these images with the replica images of intact and S1-decorated sarcomeres led us to conclude that, in intact sarcomeres, the elastic filaments were laterally associated with myosin and actin filaments in the A and I bands, respectively. Interestingly, it was shown that the elastic property of connectin filaments was not restricted by their lateral association with actin filaments in intact sarcomeres. Finally, we have proposed a new structural model of the cardiac muscle sarcomere that includes connectin filaments.

Actin Cytoskeleton↗

Histogenesis of abnormal elastic fibers in blebs and bullae of patients with spontaneous pneumothorax: ultrastructural and immunohistochemical studies.

Elastic fibers in 15 blebs and 17 bullae with spontaneous pneumothorax were studied by means of electron microscopy and light and electron microscopic immunohistochemistry for elastin and alpha 1-antitrypsin. Blebs were formed in association with focal organized alveoli, and bullae were formed in association with pulmonary emphysema. Both blebs and bullae had abnormal elastic fibers. Ultrastructurally, abnormal elastic fibers of blebs and bullae consisted of accumulated thick and fine fibers. Accumulated thick elastic fibers showed vacuolar changes and electron-dense granular deposits, and they were associated with spiraling collagen fibrils. These thick elastic fibers reacted evenly with antielastin antibody and also reacted with anti-alpha 1-antitrypsin antibody. They are thought to be degraded elastic fibers. Accumulated fine elastic fibers consisted of bundles of microfibrils and granular amorphous components, and they reacted with anti-elastin and anti-alpha 1-antitrypsin antibody. These fine elastic fibers are thought to be not only newly formed in the process of organization but also degraded. It is suggested that elastic fibers of blebs and bullae are degraded due to an imbalance between elastase and alpha 1-antitrypsin.

Adult↗

Effects of aging, menopause, and hormone replacement therapy on forearm skin elasticity in women.

OBJECTIVES: To investigate the decline per year in skin elasticity in postmenopausal women and how much hormone replacement therapy (HRT) increased elasticity over 12 months. DESIGN: Observational study of convenience sample. SETTING: The Cardiovascular Hospital of Central Japan, a cardiovascular medical center. PARTICIPANTS: One hundred seventy-six postmenopausal subjects, mean age+/-standard deviation=61.3+/-9.1. MEASUREMENTS: In Study 1, skin elasticity was measured in the right forearm using a suction device. Comparisons also were made with 45 premenopausal subjects (aged 34.5+/-9.9). Skin elasticity in the right forearm was measured using the same device at baseline and 12 months after initiation of HRT. INTERVENTIONS: In Study 2, 12 postmenopausal subjects (mean age=57.1+/-7.4, range 49-71) received conjugated equine estrogen (0.625 mg/d) in combination with medroxyprogesterone acetate (2.5 mg/d) for 12 months. RESULTS: In Study 1, significant negative correlations between skin elasticity and age and years since menopause were found (r=-0.60, P<.001 for each), as well as a 0.55% decline per year in skin elasticity. In Study 2, 12 months of HRT significantly increased skin elasticity in postmenopausal subjects, by 5.2%. CONCLUSION: After menopause, skin elasticity declined 0.55% per year; 12 months of HRT increased elasticity by 5.2%.

Adult↗

Postnatal alterations in elastic fiber organization precede resistance artery narrowing in SHR.

Resistance artery narrowing and stiffening are key elements in the pathogenesis of essential hypertension, but their origin is not completely understood. In mesenteric resistance arteries (MRA) from spontaneously hypertensive rats (SHR), we have shown that inward remodeling is associated with abnormal elastic fiber organization, leading to smaller fenestrae in the internal elastic lamina. Our current aim is to determine whether this alteration is an early event that precedes vessel narrowing, or if elastic fiber reorganization in SHR arteries occurs because of the remodeling process itself. Using MRA from 10-day-old, 30-day-old, and 6-mo-old SHR and normotensive Wistar Kyoto rats, we investigated the time course of the development of structural and mechanical alterations (pressure myography), elastic fiber organization (confocal microscopy), and amount of elastin (radioimmunoassay for desmosine) and collagen (picrosirius red). SHR MRA had an impairment of fenestrae enlargement during the first month of life. In 30-day-old SHR, smaller fenestrae and more packed elastic fibers in the internal elastic lamina were paralleled by increased wall stiffness. Collagen and elastin levels were unaltered at this age. MRA from 6-mo-old SHR also had smaller fenestrae and a denser network of adventitial elastic fibers, accompanied by increased collagen content and vessel narrowing. At this age, elastase digestion was less effective in SHR MRA, suggesting a lower susceptibility of elastic fibers to enzymatic degradation. These data suggest that abnormal elastic fiber deposition in SHR increases resistance artery stiffness at an early age, which might participate in vessel narrowing later in life.

Animals↗

Regression of hypoxic hypertension-induced changes in the elastic laminae of rat pulmonary arteries.

The elastic laminae of the pulmonary arteries (PAs) undergo a progressive structural change in hypoxic hypertension. This study focused on the reversibility of altered PA elastic laminae of the rat due to hypoxic hypertension. The structure and cross-sectional area of the PA medial elastic laminae were examined by using electron-microscopic and image-analytic approaches during recovery from 12 h and 10 days of hypoxic hypertension. At 12 h of hypoxic hypertension, the elastic laminae, which appeared homogeneous in normal control animals, were reorganized into structures composed of randomly oriented filaments, with an increase in the cross-sectional area of 70%. At 10 days of hypoxic hypertension, the elastic laminae appeared homogeneous in structure and normal in cross-sectional area despite continuous exposure to hypoxia. During recovery from 12 h of hypoxic hypertension, the medial elastic laminae regained their homogeneous structure and normal cross-sectional area after day 2. During recovery from 10 days of hypoxic hypertension, the medial elastic laminae changed from homogeneous to filamentous structures, with a progressively altered cross-sectional area that increased by 89% from recovery day 0 to day 10 and returned to the normal level on day 30. These changes were associated with alterations in the PA wall tensile stress. These results indicated that structural changes in the PA elastic laminae were reversible and that the regression process depended on the duration of exposure to hypoxia, the state of the elastic laminae, and possibly the tensile stress level in the PA wall.

Animals↗

Elastic fibers in the anulus fibrosus of the dog intervertebral disc.

A light microscopic investigation of the anulus fibrosus in cervical intervertebral discs of the dog was conducted to ascertain the arrangement and distribution of elastic fibers. Elastic fibers were observed in all lamellae of the anulus fibrosus. However, collagenous fibers were the predominant type of connective tissue fiber, and elastic fibers were randomly dispersed among them. Intralamellar (collagenous and elastic) fibers were vertically and obliquely oriented in both superficial and deep lamellae of the anulus fibrosus. All intralamellar fibers were densely and regularly arranged in superficial lamellae, but they were more loosely organized in deep lamellae. A narrow border of interlamellar, elastic fibers was observed between broader, contiguous lamellae in the superficial zone of the anulus fibrosus. Interlamellar elastic fibers wer vertically and obliquely arranged in superficial lamellae; however, they were radially oriented in deep lamellae. The deepest lamella of the anulus fibrosus consisted of a loose, three-dimensional network of intermeshing collagenous and elastic fibers. These observations suggest that elastic fibers are integral components of the articular and shock absorption mechanisms of the anulus fibrosus, and the cervical intervertebral disc of the dog is a suitable model for experimental investigation of the role of elastic fibers in intervertebral disc herniation.

Animals↗

Morphological alterations in the elastic fibers of the rabbit craniomandibular joint following experimentally induced anterior disk displacement.

Elastic fibers are important components of the connective tissue that attaches the articular disk of the craniomandibular joint (CMJ) to the skull and mandible. Biopsies of the articular disk proper and bilaminar zone (BZ) tissues from patients with anterior disk displacement (ADD) have shown previously that there is a marked loss of elastic fibers. In the present study, the effects of inducing ADD on the elastic fibers in the rabbit CMJ disk proper, BZ and condylar cartilage were investigated. The right CMJ was exposed surgically and the discal attachments were severed except for the BZ attachments. Then, the disk was displaced anteriorly and sutured to the zygomatic arch. The CMJs were removed after 1, 2 or 6 weeks and processed for histochemical demonstration of elastic fibers. The results showed osteoarthritic changes following ADD, and a significant decrease in the number of the elastic fibers in the disk proper and BZ. The remaining elastic fibers were abnormal in their appearance and orientation. In addition, ADD led to the appearance of fine elastic fibers among the chondrocytes in the hyaline cartilage of the condyle that were not present in the cartilage of the control condyle. We conclude that induced ADD can lead to a significant loss of elastic fibers in the articular disk, and result in the appearance of elastic fibers within the cartilage of the mandibular condyle.

Animals↗

Elastic skeleton of intracranial cerebral aneurysms in rats.

In an attempt to clarify the developmental mechanism of cerebral aneurysms, we studied the elastic skeleton of experimentally induced cerebral aneurysms in rats under scanning electron microscopy after hot formic acid extraction followed by freeze-drying. We produced cerebral aneurysms in 19 rats by unilaterally ligating the common carotid artery, inducing renal hypertension, and feeding beta-aminopropionitrile fumarate. The first noted change was the loss of folds protruding from the internal elastic lamina. Morphologic changes of the internal elastic lamina, considered to be primarily responsible for aneurysmal formation, occurred after the loss or disintegration of the elastic skeleton of first the intima, then the media. In large aneurysms with thick domes, we found proliferation of elastic lamellae that may reduce the risk of rupture. It seems probable that the complex elastic skeleton of the arterial wall may account for the mechanical properties of the artery and that growth of an aneurysm occurs due to disintegration of the elastic skeleton and not simply to rupture of the internal elastic lamina. We believe that such changes in the elastic skeleton are a property of the functional state of the cells that produce elastin.

Animals↗

Explaining left ventricular pressure dynamics in terms of LV passive and active elastances.

There has been much characterization of the heart as a pump by means of models based on elastance and compliance. The present paper puts forward the new concept of time-varying passive and active elastance. The biomechanical basis of cyclic elastances of the left ventricle (LV) is presented. Elastance is defined in terms of the relationship between ventricular pressure and volume as dP = EdV+ VdE, where E includes passive elastance, Ep, and active elastance, Ea. By incorporating this concept in LV models to simulate diastolic (filling) and systolic phases, a time-varying expression has been obtained for Ea, and an LV volume dependent expression has been obtained for Ep. It is proposed to use these two elastances Ea and Ep to represent the intrinsic LV properties. The active elastance, Ea, can be used to characterize the LV contractile state and represents LV pressure variation due to LV volume variation (such as during the filling and ejection phases). The passive elastance, Ep, can serve as a measure of LV resistance to filling. Furthermore, it has been demonstrated how the LV pressure dynamics (and LV pressure response to LV volume) can be explained in terms of Ea and Ep.

Blood Pressure↗

Influence of bone tissue density and elasticity on ultrasound propagation: an in vitro study.

Ultrasound (US) waves are mechanical vibrations that are applied to a material--bone tissue--in order to study its properties, that is, density, elasticity, and structure. In this study we evaluated in which way density and elasticity of the spongy bone influenced the transmission of 1.25 MHz US pulses. Twelve cylindrical specimens (diameter, 8 mm; height, 5 mm) excised from phalanxes of pig were decalcified with 0.5 M EDTA for different times (0, 2, and 5 days). During these periods, the samples underwent the following investigations: US transmission, density, and elasticity measurements. To assess the homogeneity of decalcification, the cross-sections of some samples were microradiographed. A detailed analysis of the US signal received was performed using velocity, Fourier analysis, and some parameters typical of signal processing technique. A good correlation was found between US velocity and density (r2 = 0.70); a lower correlation was found between velocity and elasticity (r2 = 0.59). If density and elasticity are considered simultaneously, the correlation with the US velocity improves significantly (r2 = 0.84). Fourier analysis enabled us to observe a shift of the main frequency toward lower values as the decalcification process advanced. We also observed that in the regressions weighted for density, US velocity correlated poorly with elasticity (r2 = 0.16), whereas signal processing parameters maintain a good correlation with elasticity (ultrasound peak amplitude [UPA], r2 = 0.48; slope, r2 = 0.62). In this study, it has been observed that when using a signal processing technique to analyze US pulses, it is possible to identify some parameters that are related in different ways to density and to elastic properties of bone. Our results show the potentiality of US technique to separate information on bone density and elasticity that X-ray-based densitometric methods do not provide.

Animals↗

Remodeling of elastic fiber components in scleroderma skin.

The skin of patients with scleroderma is characterized by an excess accumulation of collagen in the extracellular matrix of the fibrotic reticular dermis. Elastic fibers are also disrupted in this disease, however, in contrast to collagen, relatively few studies have provided information concerning the changes that occur to elastic fiber components in scleroderma. In the present study, the extracellular matrix in scleroderma skin was examined with a specific focus on the integrity of elastic fibers. Electron microscopic observations confirmed an excess of 10 nm microfibrils present in small bundles independent of amorphous elastin in the fibrotic reticular dermis. In the same area, a population of stellate-shaped fibroblasts was identified in close association with the dermal elastic fibers. In contrast to the uniform black appearance of the elastic fibers seen in the papillary dermis and in areas of the reticular dermis not infiltrated by these cells, the elastic fibers apposed to the cells were mottled in density and often almost electron-lucent. These observations suggest that the elastic fibers in the reticular dermis were being actively degraded. Results from this study provide evidence for disintegration of elastic fibers in the skin of scleroderma patients and suggest the possibility that degradation products from the elastic matrix in the diseased tissues may act as a feedback signal for increased matrix production.

Elastic Tissue↗

Morphometric analysis of elastic skin fibres from patients with: cutis laxa, anetoderma, pseudoxanthoma elasticum, and Buschke-Ollendorff and Williams-Beuren syndromes.

Computed morphometric analysis of elastic skin fibres in patients with cutis laxa, anetoderma, Williams-Beuren syndrome, pseudoxanthoma elasticum (PXE), and Buschke-Ollendorff syndrome, all clinically ascertained, was performed and compared with data obtained from healthy individuals of the same age. The diameters, area fractions (AA%) and volume fractions (VV%) occupied by pre-elastic fibres and dermal elastic fibres were determined. Irrespective of age the diameter of dermal elastic fibres followed a Gaussian distribution for all groups studied. These diameters were taken into consideration for VV% determinations. Compared with data from skin of healthy subjects of similar age range, VV% of pre-elastic fibres was significantly decreased in patients with cutis laxa, anetoderma, Williams-Beuren syndrome, and PXE and undetectable in Buschke-Ollendorff patients. VV% of dermal elastic fibres was four- to fivefold increased in Buschke-Ollendorff syndrome, two- to threefold increased in PXE skin, four- to fivefold decreased in cutis laxa and anetoderma skin and about twofold decreased in Williams-Beuren skin. The diameter of oxytalan fibres was decreased in anetoderma and Williams-Beuren syndrome while oxytalan fibre diameter was unchanged in PXE and cutis laxa. The diameter of dermal elastic fibres was increased in PXE and Buschke-Ollendorff syndrome, but was decreased in anetoderma and Williams-Beuren syndrome and unchanged in cutis laxa. We demonstrated that cutis laxa, anetoderma, Williams-Beuren syndrome, PXE, and Buschke-Ollendorff syndrome could be easily differentiated by morphometric analysis of elastic skin fibres. Thus we propose that morphometric analyses together with skin biopsies are a valuable tool for distinguishing between inherited and/or acquired skin diseases known to display alterations of elastic fibres.

Adolescent↗

Quantitative analysis of the effects of hyaluronan and aggrecan concentration and hyaluronan size on the elasticity of hyaluronan-aggrecan solutions.

We determined elasticity (G') and viscosity (G'') of various aggrecan-hyaluronan solutions using a controlled-stress rheometer with high (10 Hz) to low (0.1 Hz) frequencies. Aggrecan solution (50 mg/ml) alone showed little elasticity at any frequency, but the addition of 3300 kDa hyaluronan at 0.001-0.1 mg/ml markedly increased the elasticity, but not the viscosity, at all frequencies. Increasing hyaluronan concentration at >0.1 mg/ml did not further increase the elasticity of the aggrecan solution, and the elasticity of the aggrecan-hyaluronan complex solution reached a plateau at a 500:1 (w/w) ratio. In studies with increasing concentrations of aggrecan and a constant concentration (0.5 mg/ml) of 3300 kDa hyaluronan, aggrecan induced elasticity only at >20 mg/ml, indicating the presence of a critical concentration for elasticity. In the presence of 50 mg/ml aggrecan, 1000 kDa hyaluronan had far less effect on the elasticity of the aggrecan solution than did 3300 kDa hyaluronan. These findings suggest that only approximately 50% reduction in aggrecan concentration (<20 mg/ml), or reduced hyaluronan size (<1000 kDa)--compared with their physiological levels in young cartilage--can abolish the elastic network of the aggrecan-hyaluronan complex.

Aggrecans↗

[New method for estimating left ventricular myocardial elasticity by vibration analysis].

We solved numerical solutions of Advani-Lee's equation, which treats free vibrations of fluid-filled spherical shells, and forms the basis for non-invasive estimation of left ventricular myocardial elasticity. Numerical results showed that elasticity is approximated by E = 86.5.a2.f2 (E: elasticity (dyn/cm2), a: internal radius (cm), f: eigen-frequency (Hz)). To examine the accuracy of this theoretical equation in estimating elasticity, we made 7 spherical shells of silicone rubber, and compared the estimated elasticity by this equation with that by a standard stretch test. The elasticity calculated by Advani-Lee's equation and by stretch test proved to be nearly identical. Therefore, we concluded that we can estimate the elasticity of a spherical shell using this equation. We calculated the myocardial elasticity at the first heart sound emission in 25 normal persons with a simplified (approximated form of) Advani-Lee's equation. The mean elasticity in normal subjects was (7.04 +/- 2.46) x 10(5) dyn/cm2.

Elasticity↗

[Three-dimensional observation of elastic tissue in Bruch's membrane by formic acid extraction].

Japanese monkey and albino rabbit choroid were treated with hot-formic acid to remove the cellular elements and the extracellular matrices excluding the elastic tissues so that both three-dimensional and bird's eye views of the elastic layer of Bruch's membrane could be obtained. The only elastic layer was extracted and observed with a scanning electron microscope. Elastic layers were frequently extracted as translucent monolayered sheets with oval, sector-shaped fenestrations of varying sizes. Elastic fibers of 100-200nm in diameter crossed each other in these fenestrations. Other fine fibers connecting these elastic fibers and their sheet-like elastic plate were also observed. In Japanese monkey eyes, lattice structures formed by thick elastic fibers, 200-300nm in diameter, were conspicuous. This method proved suitable for the observation of the three-dimensional structure of the elastic layer of Bruch's membrane.

Animals↗

A selective histochemical method for the quantitative estimation of elastic fibers by computerized morphometric analysis. Effect of colchicin treatment.

A morphometric technique is reported that uses a new selective staining of the elastic system fibers in skin biopsy specimens to facilitate the quantitative evaluation of the volume fraction occupied by these elastic fibers in the tissue. The study of elastic fibers in the dermis of 30 patients, before and after six months of treatment with Colchicin, was carried out with a Quantimet 720 system. Preelastic (oxytalan and elaunin) fibers and mature elastic fibers were quantitated separately. Compared to the average volume fraction (surface occupied by the elastic fibers) before treatment with Colchicin (1.449 +/- 0.64%), the mean values after treatment were significantly increased (2.076 +/- 0.61%). The same results were found for the preelastic fibers: 0.807 +/- 0.51% before treatment and 1.025 +/- 0.54% after treatment. These results demonstrate the advantages of our monochromatic staining method for automatic quantitation of elastic fibers as well as the possibilities of the quantitative study of the elastic fibers in human dermis. This methodology should be applicable to other inherited or acquired diseases affecting skin elastic fibers as well as to other tissues containing elastic fibers.

Colchicine↗

Elastofibroma: disturbed elastic fibrillogenesis by periosteal-derived cells? An immunoelectron microscopic and in situ hybridization study.

Monospecific antibodies to elastic tissue components have been used for immunoelectron microscopy of two examples of elastofibroma. The elastic-staining fibers typically seen in these lesions exhibited a variety of morphologies with differing ratios of the amorphous and microfibrillar components usually seen in elastic fibers. The amorphous elastic material in these fibers had variable affinity for ionic stains and exhibited several substructural morphologies. Despite this, each form reacted specifically with anti-elastin antibodies. Most of the elastic fibers were associated with relatively large numbers of 12-nm diameter microfibrils that were typical of those associated with normal elastic fibers, and were specifically reactive with monospecific antibodies to microfibril-associated glycoprotein. In situ hybridization studies with a cRNA probe for human elastin confirmed that active elastin biosynthesis was occurring patchily within the lesions. The appearances and staining characteristics of the elastic tissue elements, the morphology of the cells, and the structure of the collagen fibers in these lesions were shown to have many features in common with those of normal periosteum. It is proposed that elastofibromas arise from the periosteum as a result of chronic irritation and that the different elastic fiber morphologies represent disturbances of elastic fibrillogenesis by periosteal-derived cells.

Aged↗