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Biomedical subjects

P P Purslow

Publications and source records attributed to P P Purslow.

At least 19 recordsLinked to original sources
PubMed2002

Dietary-induced changes of muscle growth rate in pigs: effects on in vivo and postmortem muscle proteolysis and meat quality.

The effects of various growth rates in pigs induced by four different feeding strategies on the activity of the calpain system and on postmortem (PM) muscle proteolysis and tenderness development were studied. An increased growth rate may be caused by an increased protein turnover, which results in up-regulated levels of proteolytic enzymes in vivo that, in turn, possibly will affect PM tenderness development. It can be hypothesized that increased proteolytic activity pre-slaughter will increase the PM tenderization rate. From postnatal d 28 to d 90 (phase 1) the pigs were divided into two groups, given either ad libitum (A) or restricted (R, 60% of ad libitum) access to feed. The two groups were then divided into two subgroups, given either restricted or ad libitum access to feed from d 91 to slaughter at d 165 (phase 2). Measurements of the activity of mu-calpain, m-calpain, and calpastatin; concentrations of total collagen and the percent of soluble collagen; and RNA, DNA, and elongation factor-2 where made at slaugther. Myofibrillar fragmentation index (MFI) was determined at slaughter and 24 h PM. Warner-Braztler shear force was determined 1 d and 4 d PM. Pigs fed restricted diets in phase 1 and fed ad libitum in phase 2 (RA pigs) had increased growth rates in the last phase compared to pigs fed ad libitum during both phase 1 and phase 2 (AA pigs). The increased growth rate (compensatory growth) was followed by an increased proteolytic potential (mu-calpain:calpastatin ratio), increased MFI values, and higher tenderization rates. There was a positive correlation between the activities of m-calpain and growth rates (r = 0.35, P = 0.03), and between RNA levels and growth rates (r = 0.43, P = 0.006). The proposed hypothesis is largely supported by the results. The activities of both mu- and m-calpain at slaughter were highest in fast-growing pigs. The calpain activity was highest in RA pigs, which in turn also had the fastest growth rates prior tslaughter among the four groups. This implies that the synthesis of these enzymes was up-regulated during the second feeding period to a larger extent in RA pigs. The proteolytic potential and the MFI values indicate that the up-regulated in vivo calpain activity had an effect on PM protein degradation, which also is supported by the higher tenderization rate in RA pigs.

Animal Feed↗
PubMed2001

Role of Ca(2+) for the mechanical properties of fibrillin.

Fibrillin-rich microfibrils are important structural elements widespread throughout connective tissues. Genetic defects identified in the Ca(2+) binding sites of fibrillin have severe effects and in addition Ca(2+) has a marked effect on the microfibrillar structure. We have studied the role of Ca(2+) on the mechanical behavior of fibrillin-rich microfibrils using the micro-needle technique. We find that Ca(2+)-depletion results in a 50% decrease in rest length and reduces the stiffness of fibrillin-rich microfibrils. At high strain, irreversible damage occurs. This behavior is consistent with Ca(2+) stabilization of interactions between consecutive EGF-like domains and breakdown in the quaternary structure upon over-extension.

Animals↗
PubMed2001

Fibrillin-rich microfibrils of the extracellular matrix: ultrastructure and assembly.

Fibrillin-rich microfibrils are a unique class of extensible connective tissue macromolecules. Their critical contribution to the establishment and maintenance of diverse extracellular matrices was underlined by the linkage of their principal structural component fibrillin to Marfan syndrome, a heritable connective tissue disorder with pleiotropic manifestations. Microscopy and preparative techniques have contributed substantially to the understanding of microfibril structure and function. The supramolecular organisation of microfibrillar assemblies in tissues has been examined by tissue sectioning and X-ray diffraction methods. Published findings are discussed and new information reported on the organisation of microfibrils in the ciliary zonular fibrils by environmental scanning electron microscopy. This review summarises microscopy and X-ray diffraction studies that are informing current understanding of the ultrastructure of fibrillin-rich microfibrils.

Ectopia Lentis↗
PubMed2001

Development of components of the extracellular matrix, basal lamina and sarcomere in chick quadriceps and pectoralis muscles.

1. The purpose of this study was to investigate differences in the development of components of the cell/matrix linkage in two functionally different muscle types: the pectoralis muscle, a major locomotory muscle in birds but not particularly functional in chickens, and the quadriceps muscle, a smaller and more functionally active muscle in the chicken. 2. The development of the extracellular matrix, basal lamina and sarcomere in the pectoralis and quadriceps muscles in chick embryos was examined biochemically to determine differences in the rate of development between these two muscles. Samples of these muscle types were dissected out from chick embryos from embryonic day 10 until 8 weeks post hatch. 3. Using SDS-PAGE electrophoresis and western blotting with antibodies against sarcomeric actin, laminin and collagens I, III and IV, it was apparent that muscle development begins earlier in the quadriceps muscle than in the pectoralis, and that late in the developmental process (d 18) both muscle types were well differentiated. The final concentration of collagens in the mature muscle remained higher in the quadriceps than in the pectoralis muscle. 4. The onset of development of the extracellular matrix, basal lamina and sarcomere was earlier in the quadriceps than the pectoralis, which could have functional implications for these muscles as a whole.

Actins↗
PubMed2000

Epinephrine upregulates calpain activity in cultured C2C12 muscle cells.

C2C12 cells were grown to confluence at 37 degrees C under a continuous 5% CO(2) stream and myotube formation was stimulated. The cultures were then incubated with or without 2 microg/mL epinephrine for 18 h prior to harvesting and calpain extraction. Epinephrine treatment resulted in a three-fold increase in extractable mu-calpain activity (P < 0.05), a three-fold increase in extractable m-calpain activity (P < 0.05), a 36% increase in calpastatin activity (P < 0.001), and a 16% decrease (P < 0.05) in the total protein content in the C2C12 cell homogenate. These results suggest that calpains may play a role in protein metabolism and that the hormone epinephrine may be directly involved in the regulation of their cellular expression.

Animals↗
PubMed2000

Age related compliance of the lamina cribrosa in human eyes.

AIMS: To investigate changes in the mechanical compliance of ex vivo human lamina cribrosa with age. METHODS: A laser scanning confocal microscope was used to image the surface of the fluorescently labelled lamina cribrosa in cadaver eyes. A method was developed to determine changes in the volume and strain of the lamina cribrosa created by increases in pressure. The ability of the lamina cribrosa to reverse its deformation on removal of pressure was also measured. RESULTS: Volume and strain measurements both demonstrated that the lamina cribrosa increased in stiffness with age and the level of pressure applied. The ability of the lamina cribrosa to regain its original shape and size on removal of pressure appeared to decrease with age, demonstrating an age related decrease in resilience of the lamina cribrosa. CONCLUSIONS: The mechanical compliance of the human lamina cribrosa decreased with age. Misalignment of compliant cribriform plates in a young eye may exert a lesser stress on nerve axons, than that exerted by the rigid plates of an elderly lamina cribrosa. The resilience of the lamina cribrosa also decreased with age, suggesting an increased susceptibility to plastic flow and permanent deformation. Such changes may be of importance in the explanation of age related optic neuropathy in primary open angle glaucoma.

Adolescent↗
PubMed2000

Differentiation of myoblasts in serum-free media: effects of modified media are cell line-specific.

Myoblast cell lines are grown and differentiated readily in cell culture. Two cell lines typically used for investigating the growth and differentiation of muscle are the mouse cell line C2C12 and the rat cell line L6. The differentiation of these cells in vitro requires a switch from a serum-rich medium to a less rich medium after the cells have reached confluence. Since the components present in serum are not well characterized, the use of a better defined medium for these studies was investigated. C2C12 and L6 myoblasts were differentiated in both serum-containing and serum-free media. The differentiation state of these cultures was then tested both microscopically and biochemically. Cultures were checked for myotube formation, the activity of creatine phosphokinase and the presence of sarcomeric actin. In C2C12 cells, the extent of differentiation was greater in the serum-free than in the serum-containing system. In both media types, the C2C12 cells produced sarcomeric actin, showing the presence of sarcomere structure in the myotubes. In L6 cells, however, myotubes were readily formed in medium containing 2% horse serum, but not in the serum-free system. In addition, the ability of C2C12 cells to differentiate on substrates coated with extracellular matrix proteins was shown to be media-dependent. The presence of extracellular matrix proteins did not enable L6 cells to form myotubes when cultured in serum-free media. Primary cultures of chick myoblasts were able to differentiate in both media tested, with Dulbecco's modified Eagle medium containing horse serum being a more efficient medium for cell fusion. This study shows a divergence in muscle cell line responses in three cell lines, two of which are typically used as 'model systems' for understanding muscle growth and development.

Actins↗
PubMed1999

The supramolecular organisation of fibrillin-rich microfibrils determines the mechanical properties of bovine zonular filaments.

The zonular filaments from the eyes of cows are rich in microfibrils containing fibrillin. Tensile tests, stress-relaxation tests and X-ray diffraction studies were used to study the relationship between the mechanical behaviour of zonular filaments and the molecular packing and structure of the fibrillin-rich microfibrils. Zonular filaments show a non-linear (J-shaped) stress-strain curve and appreciable stress-relaxation. It is proposed that the non-linear properties are due to local variations in waviness in the microfibrils or assemblies of microfibrils, which straighten out and become more regularly aligned with strain. Previous and current X-ray diffraction results consistently show a partial ordering of microfibrils in zonular filaments into staggered aggregates which become more ordered and laterally aligned on stretching. Although the removal and re-addition of Ca(2+) is known to change the molecular structure of fibrillin, no effect was observed on the tensile properties of the zonular filaments. It is hypothesised that strain-induced deformation in the supramolecular aggregate packing may not be Ca(2+)-sensitive but could dominate the mechanical behaviour of microfibrillar arrays in zonular filaments.

Animals↗
PubMed1998

Calcium determines the supramolecular organization of fibrillin-rich microfibrils.

Microfibrils are ubiquitous fibrillin-rich polymers that are thought to provide long-range elasticity to extracellular matrices, including the zonular filaments of mammalian eyes. X-ray diffraction of hydrated bovine zonular filaments demonstrated meridional diffraction peaks indexing on a fundamental axial periodicity (D) of approximately 56 nm. A Ca2+-induced reversible change in the intensities of the meridional Bragg peaks indicated that supramolecular rearrangements occurred in response to altered concentrations of free Ca2+. In the presence of Ca2+, the dominant diffracting subspecies were microfibrils aligned in an axial 0.33-D stagger. The removal of Ca2+ caused an enhanced regularity in molecular spacing of individual microfibrils, and the contribution from microfibrils not involved in staggered arrays became more dominant. Scanning transmission electron microscopy of isolated microfibrils revealed that Ca2+ removal or addition caused significant, reversible changes in microfibril mass distribution and periodicity. These results were consistent with evidence from x-ray diffraction. Simulated meridional x-ray diffraction profiles and analyses of isolated Ca2+-containing, staggered microfibrillar arrays were used to interpret the effects of Ca2+. These observations highlight the importance of Ca2+ to microfibrils and microfibrillar arrays in vivo.

Animals↗
PubMed1998

X-Ray diffraction studies of fibrillin-rich microfibrils: effects of tissue extension on axial and lateral packing.

X-ray diffraction of hydrated fibrillin rich microfibrils, in the form of zonular filaments from bovine eyes, demonstrated meridional diffraction peaks indexing on a fundamental periodicity of approximately 56 nm in the relaxed state. The effect of sample extensions of up to 50% in length produced an increase in the axial periodicity of only 4% as judged by alteration of the diffraction peak position of the third meridional order. This effect was shown to be reversible. Further extension to 100% of the tissue rest length caused extensive deterioration in the quality of the diffraction and resulted in a more complex meridional diffraction series, where the fundamental axial periodicity also changed to a length of approximately 80 nm. The fibrillin diffraction image also contains an equatorial diffraction peak that is enhanced upon tissue extension. The measurement of the molecular spacing from the equatorial diffraction profile indicated that the closest approach of molecules gave a broad interference peak of spacing 28 nm, this is nearly twice the molecular diameter as estimated from electron microscopy of dehydrated samples.

Animals↗
PubMed1998

Collagen orientation and molecular spacing during creep and stress-relaxation in soft connective tissues.

Collagen fibres form cross-helical, cross-ply or quasi-random feltworks in extensible connective tissues; strain-induced reorientation of these networks gives rise to the non-linear mechanical properties of connective tissue at finite strains. Such tissues are also generally viscoelastic (i.e. display time-dependent properties). The hypothesis that time-dependent reorientation of collagen fibres is responsible for the viscoelasticity of such tissues is examined here using time-resolved X-ray diffraction measurements during stress-relaxation and creep transients applied to rat skin and bovine intramuscular connective tissue. Differences in the intensity and angular orientation of the third and fifth orders of the 67 nm meridional D-spacing of collagen molecules were shown before and after the application of loads or displacements. However, no changes in the D-spacing or angular orientation of collagen occurred during the time course of either stress-relaxation or creep in both tissues. This indicates that collagen fibre reorientation is not a primary source of their viscoelastic properties. The non-linear (strain-dependent) nature of the stress-relaxation response in these tissues suggests that relaxation processes within the collagen fibres or at the fibre-matrix interface may be responsible for their viscoelastic nature.

Animals↗
PubMed1997

Fibrillin-rich microfibrils: an X-ray diffraction study of the fundamental axial periodicity.

Microfibrils are ubiquitous matrix polymers which are thought to provide elastic properties in all extracellular matrix structures. The major component of the elastic microfibrils is the protein fibrillin; its molecular structure is unknown. In electron microscopy, microfibrils appear as beaded structures exhibiting a variable periodicity, indicating that they may be elastomeric. The X-ray diffraction of fibrillin-rich microfibrils in the form of zonular filaments from bovine eyes exhibits meridional diffraction peaks indexing on a fundamental periodicity of 55 nm in the relaxed state. The application of a 40% extension produced a lengthening of the periodicity by 3% as judged by alteration of the D spacing of the principal peaks. This effect was shown to be reversible. Changes in the periodicity of the meridional reflections indicate changes in the fundamental structure of the microfilaments, but cannot account for all long range elastomeric properties of fibrillin-containing microfibrils.

Actin Cytoskeleton↗
PubMed1997

Biomechanical and biochemical study of a standardized wound healing model.

Standardized protocols were developed for use in a detailed investigation into the biomechanical and biochemical properties of a dermal wound healing model in the rat. The use of a rapid freezing method at -80 degrees C minimized the detrimental effects of freezing on the biomechanical properties of the tissue and also allowed for convenient inter-laboratory collaboration to be performed. The methodology described allowed for the simultaneous and reproducible measurement of tensile strength, collagen cross-linking and proteolytic enzyme activity. Increases in the tensile properties of the tissue with time were consistent with an active process of remodelling process as indicated by changes in the cross-link and enzyme profiles. Initially the granulation tissue was comparatively rich in the keto-imine cross-link hydroxylysino-keto-norleucine, which was later replaced by the aldimine cross-link dehydro-hydroxy-lysinonorleucine. The mature cross-link histidino-hydroxy-lysinonorleucine was not observed within the granulation tissue at any stage and was also absent in aged control skin. A peak of matrix metalloproteinase-9 activity was observed at early timepoints (48 hr) and then decreased rapidly to normal levels and is consistent with an acute inflammatory response. In contrast matrix metalloproteinase-2 activity peaked later (3 days) and then decreased gradually, consistent with its role as one of the predominant enzymes involved in the remodelling process. The results described validate the animal model used and emphasize its potential for use in combined biomechanical and biochemical studies of acute wound healing.

Animals↗
PubMed1996

Ocular elasticity. Is engineering stiffness a more useful characterization parameter than ocular rigidity?

PURPOSE: The purpose of this study is to present an analysis of the pressure-volume relation of the eye in basic engineering terms, so as to characterize the deformability of the ocular shell based on its intrinsic stiffness (Young modulus) and morphology, as opposed to the empirical measure of ocular rigidity. METHODS: Starting from the structural mechanics equations describing the stress of spherical thin-walled vessels, the differential equation governing the eye pressure-volume relation is derived. This analysis, which is more rigorous than previously published derivations, assumes that the ocular shell has a Poisson ratio of 0.5. This assumption is experimentally confirmed by ultrasonic measurements of changes in bovine corneal thickness with intraocular pressure. RESULTS: Even with a number of simplifying assumptions, this basic analysis yields a complex result, showing that the Young modulus of the ocular shell material increases rapidly with distension of the eye, and is approximately proportional to the fourth power of the ocular shell radius. CONCLUSION: Due to the complexity of the phenomenon, engineering analysis does not lead to a simple picture of pressure-volume relation of the eye. However, it does explicitly separate the material properties of the ocular shell from morphologic contributions to pressure-volume relation of the eye. This approach allows pathologic changes in the pressure-volume relation of the eye to be related more easily to the fundamental structural mechanisms governing the nonlinear mechanical properties of ocular shell materials.

Animals↗
PubMed1994

The morphology and mechanical properties of endomysium in series-fibred muscles: variations with muscle length.

In the series-fibred muscle architecture commonly found in large muscles of mammals and birds, the intrafasciculary-terminating muscle fibres have no direct tendinous attachments. Contractile force produced in these fibres must be transmitted between adjacent muscle fibres via the endomysial connective tissue which separates them. The endomysium is thus an essential mechanical component in such muscles. Studies of motor end-plate banding patterns and the frequent occurrence of tapering ends of fibres within the fascicles of the bovine sternomandibularis muscle show it to be a series-fibred muscle. Sodium hydroxide digestion of fixed samples of this muscle to remove the myofibrillar apparatus revealed the endomysium to be a disordered planar network of mainly curvilinear collagen fibrils. The orientation distribution of the collagen fibrils in the endomysial network was measured by image analysis of scanning electron micrographs. Analysis of endomysial preparations from muscle fixed at sarcomere lengths between 1-4 microns showed that the orientation distribution of collagen fibrils is quantitatively related to muscle length. At rest sarcomere length the collagen fibril network is not completely random, but has a slight circumferential bias. The orientation distribution shows a progressive shift towards the circumferential direction at short sarcomere lengths and towards the longitudinal direction at long sarcomere lengths. The relationship between the number-weighted mean collagen orientation and sarcomere length was compared to two geometric models of network behaviour, the isoareal and constant shape models. Both fitted the data reasonably, although the constant shape model described the rate of change of mean orientation more closely. From fibrous composites theory, the reinforcement efficiency factor, eta, was calculated from the measured collagen fibril orientation distributions. These calculations predict a non-linearly increasing longitudinal tensile modulus for the endomysium with increasing sarcomere length, in agreement with its known non-linear properties, but confirm that the tensile properties of the endomysium are unsuitable for transmission of tensile force from muscle fibres contracting near rest length. This reinforces a previous interpretation that contractile force is transmitted between neighbouring muscle fibres by trans-laminar shear through the endomysium rather than by in-plane tension.

Animals↗
PubMed1992

Functional morphology of the endomysium in series fibered muscles.

Many skeletal muscles, including the feline biceps femoris, are composed of short, tapered myofibers arranged in an overlapping longitudinal series. The endomysium of such muscles transfers tension between overlapping myofibers, and is thus an elastic element in series with them. The endomysium of the cat biceps femoris contains curvilinear collagen fibrils in an approximately isotropic (random) array. The collagen fibrils undergo only a modest reorientation as the myofibers shorten or lengthen within the physiological range. A geometrical model predicts no change in the thickness of the endomysium on changing muscle fiber length and quantifies the expected collagen fibril reorientation in the endomysium as a function of muscle extension. It is also demonstrated that a high proportion of the collagen fibrils will be curvilinear at all sarcomere lengths. The organization of endomysial collagen is appropriate for the transfer of loads between myofibers by means of shear.

Animals↗
PubMed1989

Strain-induced reorientation of an intramuscular connective tissue network: implications for passive muscle elasticity.

The most abundant intramuscular connective tissue component, the perimysium, of bovine M. sternomandibularis muscle was shown to be a crossed-ply arrangement of crimped collagen fibres which reorientate and decrimp on changing muscle fibre sarcomere length. Reorientation of perimysial strands was observed by light microscopy and identification of these strands as collagen fibres was confirmed by high-angle X-ray diffraction. Mean collagen fibre direction with respect to the muscle fibres ranged from approximately 80 degrees at sarcomere length = 1.1 micron to approximately 20 degrees at 3.9 microns. This behaviour was well described by a model of a crimped planar network surrounding a muscle fibre bundle of constant volume but varying length. Modelling of the mechanical properties of the perimysium at different sarcomere lengths produced a load-sarcomere length curve which was in good agreement with the passive elastic properties of the muscle, especially at long sarcomere lengths. It is concluded that the role of the perimysial collagen network is to prevent over-stretching of the muscle fibre bundles.

Animals↗