Effects of Reduced Joint Mobility on Sarcomere Length,Collagen Fibril Arrangement in the Endomysium,and Hyaluronan in Rat Soleus Muscle

Immobilization is often associated with a decrease in muscle elasticity. This condition is called muscle contracture, but the mechanism is not yet clear. We examined changes in ankle joint mobility, sarcomere length, collagen fibril arrangement in the endomysium, and hyaluronic acid (HYA) in muscular tissue 1, 2, 4, 8, and 12 weeks after immobilization of rat soleus muscles in shortened position. Ankle joint mobility decreased with the duration of immobilization. Sarcomere length had shortened 1 week after immobilization, but did not show further change 2, 4, 8, and 12 weeks after immobilization. Collagen fibril arrangement in the endomysium 1 and 2 weeks after immobilization was longitudinal to the axis of the muscle fibers, whereas 4, 8, and 12 weeks after immobilization it was circumferential. HYA in muscular tissue had increased 1 week after immobilization but remained at the same level at weeks 2, 4, 8, and 12. Histochemically, HYA in the endomysium of immobilized muscular tissue was stained more strongly and widely than that in the control tissue. Increased HYA in muscular tissue may induce muscle stiffness, but the significance of how HYA is related to the mechanism of muscle contracture was not clear. The findings suggest that muscle contracture started 1 week after immobilization and increased with the length of immobilization. Consequently, muscle contracture is affected by the shortening muscle fibers during the early stage of immobilization, after which the collagen adapts by the fibril arrangement in the endomysium becoming more circumferential. This change in collagen fibril arrangement may cause advanced muscle contracture in the late stage of immobilization.     

Heterogeneity of mean sarcomere length in different fibres: effects on length range of active force production in rat muscle

Isometric active length-force characteristics of rat semimembranosus lateralis muscles [rats,n = 6, body mass = 292 (SD 9) g] were recorded. Muscles were photographed during the force plateaus of the tetani to determine lengths of proximal, intermediate and distal fibres. From the mean number of sarcomeres in series in those fibres, mean sarcomere length at different muscle lengths was calculated. The heterogeneity of mean sarcomere lengths for each muscle was quantified according to the coefficient of variation of sarcomere lengths at muscle optimal length. Absolute muscle length changes were equal to fibre length changes. At all muscle lengths, mean sarcomere lengths in the distal fibres were significantly greater than those in proximal and intermediate fibres. The heterogeneity of mean sarcomere lengths augmented the length range of active force production between muscle optimal length and active slack length by about 38% to 43%. We concluded that in a muscle with a low degree of pennation, the heterogeneity of mean sarcomere lengths should be considered as a substantial contributor to the length range over which active force can be produced. In our experiment, the length ranges of active force production between optimal and slack length showed considerable differences (range 18.7–24.9 mm). The coefficient of correlation between length ranges and mean number of sarcomeres in series in various muscle regions was extremely low (r = –0.14) and not significant. The coefficient of correlation between length ranges and heterogeneity was high (r = 0.88) and significant. These data would suggest that muscles with a similar number of sarcomeres in series can exhibit quite different functional characteristics.     

Structural arrangement of collagen fibrils in the periarterial connective tissue of the kidney: their functional relevance as a structural stabilizer against arterial pressure

Periarterial connective tissue with a moderate amount of collagen fibrils is known to be a specialized domain in the renal interstitium. This study aimed to clarify the microscopic architecture of the periarterial connective tissue as a mechanical supportive structure of the intrarenal arteries. Transmission and scanning electron microscopy revealed two populations of collagen fibrils in the periarterial connective tissue. The major one was composed of many bundles of collagen fibrils running in longitudinal directions, whereas the minor one was represented by a few circumferential bundles adjacent to the smooth muscles. The amount of collagen fibrils was obviously variable and correlated with the arterial caliber. The correlation between abundance of collagen fibrils and the arterial caliber was confirmed by morphometric analysis of the collagen fibril area per arterial perimeter on electron micrographs. The size of individual collagen fibrils was measured in periarterial connective tissue of arteries with various calibers. A positive correlation between the diameter of collagen fibrils and arterial caliber was confirmed, indicating the supportive function of collagen fibrils in the periarterial connective tissue. The accumulated morphological findings supported the hypothesis that the collagen fibrils in the periarterial connective tissue develop longitudinal tension with their tensile strength, whereas the smooth muscle cells in the media develop circumferential tension with active regulation of contracting force.     

The descending limb of the sarcomere length-force relation in single muscle fibres of the frog

Summary Single muscle fibres, isolated from the tibialis anterior muscle of the frog, were used to study intersarcomere dynamics during muscle-isometric (fixed-end) tetani at long sarcomere lengths. Sarcomere length was measured by an online laser diffraction technique. On the descending limb of the length-force relation, the slow rise of force (creep) was always associated with changes in sarcomere length. Sarcomeres at the ends of the fibres shortened, while those of the central 90% of the fibre length were stretched. Fibres were found to have a range of passive length-force curves, those with high resting forces developed little creep force, while low resting force fibres developed substantial creep, resulting in a fixed-end sarcomere length-force relation which deviated greatly from that expected from crossbridge theory. These differences in creep force can be qualitatively accounted for by differences in sarcomere dynamics. The simultaneous measurement of force and sarcomere length during force development allows the construction of a sarcomere-isometric length-force curve from minima in the sarcomere length record. Force declined linearly from a plateau at 2.2 µm to zero at a sarcomere length close to 3.65 µm. The online, diffraction-derived sarcomere length was used in a feedback loop to clamp sarcomere length in short (100–200 µm) segments of fibres. A length-force curve constructed from sarcomere length-clamped tetani shows a linear decline in force from a plateau at 2.2 µm to zero at a sarcomere length of 3.65 µm.     

Mechanisms causing effects of muscle position on proximo-distal muscle force differences in extra-muscular myofascial force transmission

Certain recent studies showed that extra-muscular myofascial force transmission affects the length-force characteristics of rat extensor digitorium longus (EDL) muscle significantly after distal or proximal lengthening. This suggested that the relative position of a muscle with respect to its surrounding connective tissues is a co-determinant of muscle force in addition to muscle length, and indicated major effects on muscular mechanics. The specific goal of the present study is to investigate such effects by studying: (1) distributions of lengths of sarcomeres within muscle fibres and (2) the relative contributions of muscle fibres and the extra-cellular matrix to muscle total force, using a finite element model. The length of the muscle modelled was kept constant at a high and at a low muscle length whereas the relative position of the muscle was altered exclusively. For both muscle lengths, the forces exerted at distal and proximal tendons were unequal at almost all muscle relative positions. The proximo-distal force difference was enhanced as the muscle was repositioned away from its reference position. This confirmed the role of relative position of a muscle as a co-determinant of muscle force. At higher muscle lengths, distributions of lengths of sarcomeres arranged in series within muscle fibres were substantial. The force transmitted by the muscles' extra-cellular matrix comprised a sizable part of muscle total force. At lower muscle lengths distribution of sarcomere lengths was relatively limited indicating that the extra-cellular matrix is bearing the extra-muscular force. However, minor sarcomere length changes were shown to accumulate to sizable effects on the summed forces exerted by the muscle fibres. In addition, the extra-muscular load was shown to manipulate the force exerted by the extra-cellular matrix. We conclude that the relative position of a muscle has substantial effects on intra-muscular mechanics and the importance of the role of the extra-cellular matrix in determining the proximo-distal force differences is comparable to that of the intra-cellular domain.     

Organization and distribution of intramuscular connective tissue in normal and immobilized skeletal muscles

Collagen fiber network is a major contributor to the coherence and tensile strength of normal skeletal muscle. Despite the well-recognized importance of the intramuscular connective tissue to the normal integrity and function of the skeletal muscle, the specific architecture including the location and three-dimensional orientation of the intramuscular connective tissue within the muscle tissue is poorly described. The structure of the intramuscular connective tissue was studied by immunohistochemistry, polarization microscopy (the crimp length and angle of the collagen fibers) and scanning electron microscope (SEM) in rat skeletal muscles (gastrocnemius, soleus and tibialis anterior) in normal situation and after 3 weeks of disuse (immobilization). Three separate networks of collagen fibers were distinguished by SEM in the normal endomysium; fibers running longitudinally on the surface of the muscle fibers (the main collagen orientation), fibers running perpendicularly to the long axis of the muscle fibers and having contacts with adjacent muscle fibers, and fibers attached to the intramuscular nerves and arteries. Similarly, the SEM analysis also disclosed three distinct collagen fiber networks running in different directions in the perimysium, but, contrary to the endomysium, the main fiber orientation could not be established. Immobilization resulted in a marked increase in the endo- and perimysial connective tissue, the majority of the increased endomysial collagen being deposited directly on the sarcolemma of the muscle cells. Immobilization also resulted in substantial increase in the number of perpendicularly oriented collagen fibers with contacts to two adjacent muscle fibers in the endomysium. Further, immobilization clearly disturbed the normal structure of the endomysium making it impossible to distinguish the various networks of fibers from each other. In the perimysium, immobilization-induced changes were similar, the number of longitudinally oriented collagen fibers was increased, the connective tissue was very dense, the number of irregularly oriented collagen fibers was markedly increased, and consequently, the different networks of collagen fibers could not be distinguished from each other. Of the three studied intact muscles, the crimp angle of the collagen fibers was lowest in the soleus and highest in the gastrocnemius muscle, and the crimp angle decreased over 10% in all muscles after the immobilization-period. Altogether, the above described quantitative and qualitative changes in the intramuscular connective tissue are likely to contribute to the deteriorated function and biomechanical properties of the immobilized skeletal muscle.     

The microtexture of amnion and chorion connective tissue

On microscopical, stereo-scan and TEM observations the microtexture of the connective tissue of amnion and chorion have been analysed. The "compact layer" of the amnion-connective tissue consists of single collagen fibrils, which are arranged felt-like in parallel layers. In the "fibroblast layer" the fibrils are mostly arranged in bundles, forming a network. Single fibrils and bundles of fibrils in both layers are arranged surface parallel. The superficial layer of the chorion's connective tissue is predominated by wave-like bundles of fibrils in parallel and latticed arrangements, which are consolidated by transversely and diagonally running fibres. In the inferior layer the fibres leave their formation, run diagonally in the direction of the trophoblast and interweave with each other to a mat of fibrils at the border to the trophoblast. The tensile strength of the amnion connective tissue is due to the felt-like fibril texture of the "compact-layer" and the special arrangement of surface-parallel collagen fibres and fibrils.     

Aligned fibrillar collagen matrices obtained by shear flow deposition

Here we present a new technique to generate surface-bound collagen I fibril matrices with differing structural characteristics. Aligned collagen fibrils were deposited on planar substrates from collagen solutions streaming through a microfluidic channel system. Collagen solution concentration, degree of gelation, shear rate and pre-coating of the substrate were demonstrated to determine the orientation and density of the immobilized fibrils. The obtained matrices were imaged using confocal reflection microscopy and atomic force microscopy. Image analysis techniques were applied to evaluate collagen fibril orientation and coverage. As expected, the degree of collagen fibril orientation increased with increasing flow rates of the solution while the matrix density increased at higher collagen solution concentrations and on hydrophobic polymer pre-coatings. Additionally, length of the immobilized collagen fibrils increased with increasing solution concentration and gelation time.     

Force-calcium relations in skinned twitch and slow-tonic frog muscle fibres have similar sarcomere length dependencies

Summary The sarcomere length (SL) dependence of the calcium sensitivity of force was measured in skinned single twitch and slow-tonic muscle fibres from frog and toad. Twitch and slow-tonic fibres were characterized by location, appearance, physiological response to calcium and by protein band patterns from sodium-dodecyl-sulphate polyacrylamide gel electrophoresis (SDS-PAGE). Force-calcium relations were determined for each fibre type at two sarcomere lengths, 2.4 and 3.1 m. Bathing solution ionic strength (IS) was 200 mm and solution pH was 7.0, 6.0 or 5.5; experiments were also done at IS=120 mm and pH 7.0. At all pHs and ionic strengths tested, slow-tonic fibres exhibited a slower time course of force development and were more sensitive to calcium than were twitch fibres. Lowering IS increased calcium sensitivity and lowering pH decreased calcium sensitivity in both fibre types. Increasing SL increased the calcium sensitivity of force in both twitch and slow-tonic fibres at pH 7.0 and at both 200 and 120 mm IS. Lowering pH caused a decrease in the length dependence of calcium sensitivity of both fibre types; at pH 5.5 the calcium sensitivity of force in slow-tonic fibres exhibited a slight decrease with increasing SL.     

Active tension generation in isolated skeletal myofibrils

Summary Single or double myofibrils isolated from rabbit psoas muscle were suspended between a fine needle and an optical force transducer. By using a photodiode array, the length of every sarcomere along the specimen could be measured. Relaxed specimens exhibited uniform sarcomere lengths and their passive length-tension curve was comparable to that of larger specimens. Most specimens could be activated and relaxed four to five times before active force levels began to decline; some specimens lasted for 10–15 activation cycles. Active tension (20–22°C) was reproducible from contraction to contraction. The contractile response was dependent on initial sarcomere length. If initially activated at sarcomere lengths of 2.7 m, one group of sarcomeres usually shortened to sarcomere lengths of 1.8–2.0 m, while the remaining sarcomeres were stretched to longer lengths. Myofibrils that were carefully activated at shorter initial sarcomere lengths usually contracted homogeneously. Both homogeneous and inhomogeneous contractions produced high levels of active tension. Calcium sensitivity was found to be comparable to that in larger preparations; myofibrils immersed in pCa 6.0 solution generated 30% of maximal tension, while pCa 5.5–4.5 resulted in full activation. Active tension at full overlap of thick and thin filaments ranged from 0.34 to 0.94 N mm^-2 (mean of 0.59 N mm^-2±0.13 sd. n=65). Even allowing for a maximum of 20% nonmyofibrillar space in skinned or intact muscle fibres, the mean tension generated by isolated myofibrils per cross-sectional area is higher than by fibre preparations.     

Three-dimensional ultrastructure reconstruction of tendinous components at the bifurcation of the bovine superficial digital flexor tendon using array and STEM tomographies

Tendons transmit force from muscle to bone for joint movement. Tenocytes are a specialized type of fibroblast that produces collagen fibrils in tendons. Their cytoplasmic processes form a network surrounding collagen fibrils to define a collagen fibre. Glycosaminoglycan (GAG) chains link collagen fibrils and adhere at the D-band of the collagen fibril. In this study, we used array and scanning transmission electron microscope (STEM) tomographies to reconstruct the three-dimensional ultrastructure of tenocytes, collagen fibres, collagen fibrils and GAG chains at the bifurcation of the bovine hindlimb superficial digital flexor tendon (SDFT). Collagen fibrils comprising a collagen fibre were not aligned uniformly and had at least two running directions. Spindle-shaped tenocytes were arranged along the long axis of a plurality of collagen fibres, where two groups of collagen fibrils with oblique directions to each other exhibited an oblique overlap of the two collagen fibril layers. Collagen fibrils with different running directions were observed in separating layers of about 300 nm in thickness and had diameters of 0–200 nm. About 40% of all collagen fibrils had a peak in the range of 20–40 nm. STEM analysis of the same site where the crossing of collagen fibres was observed by transmission electron microscopy demonstrated the outline of collagen fibrils with a clear D-banding pattern at a regular interval. Collagen fibrils were reconstructed three-dimensionally using continuous images acquired by STEM tomography, which confirmed that the collagen fibrils at the crossing sites did not orientate in layers, but were woven one by one. Higher magnification observation of GAG chains attached between the crossing collagen fibrils revealed numerous GAG chains arranged either vertically or obliquely on collagen fibrils. Furthermore, GAG chains at the cross of collagen fibrils connected the closest D-bands. GAG chains are thought to be universally present between collagen fibrils of the tendon. These observations by array and STEM tomographies increase our knowledge of the anatomy in the bifurcation of the bovine hindlimb SDFT and demonstrate the utility of these new imaging technologies.     

An unusual Z-system in the obliquely striated muscles of crinoids: three-dimensional structure and computer simulations

Summary The peculiar functional structure of the Z-line in the obliquely striated muscles of some feather stars is described. It is known that cross-striated muscles are characterized by linear and continuous Z-bands, and obliquely striated muscles by disconnected, obliquely aligned Z-elements. Owing to this discontinuous organization, the sarcomere can perform wide active lengthenings, shortenings, and even super-elongations in the helical fibres. In contrast, the obliquely striated fibres of crinoids show markedly continuous and homogeneous oblique Z-lines; such a structure is not compatible with super-performances like sliding and shearing of the sarcomere elements, but instead could allow functions comparable to those characteristic of a cross-striated muscle (quick, short movements, mechanically amplifiable by bone levers). This odd situation, only interpretable in terms of evolutionary constraint, could be considered opposite and symmetrical to that of cross-striated super-contracting muscles, where the Z-line is exceptionally fragmented to allow the sarcomere to super-contract.The possible architecture of a significant parameter such as the Z-line, which determines muscle fibre potential capacities, is analysed in detail: (1) through qualitative-quantitative evaluation of electron micrographs, supported by statistical analysis of the data; and (2) bycomputer simulations. The data obtained suggest that the most realistic conformation of the whole Z-complex in these muscles consists of a multiple system of continuous, ribbon-like helical planes running in parallel along the fibre from end to end and regularly cutting it with a constant thickness. The proposed model seems morphologically compatible with the experimentally verified situations and functionally compatible with the mechanical requirements for a normal contraction and for a balanced distribution of the involved strengths.     

Myofascial force transmission via extramuscular pathways occurs between antagonistic muscles

Most often muscles (as organs) are viewed as independent actuators. To test if this is true for antagonistic muscles, force was measured simultaneously at: (1) the proximal and distal tendons of the extensor digitorum muscle (EDL) to quantify any proximo-distal force differences, as an indicator of myofascial force transmission, (2) at the distal tendons of the whole antagonistic peroneal muscle group (PER) to test if effects of EDL length changes are present and (3) at the proximal end of the tibia to test if myofascially transmitted force is exerted there. EDL length was manipulated either at the proximal or distal tendons. This way equal EDL lengths are attained at two different positions of the muscle with respect to the tibia and antagonistic muscles. Despite its relatively small size, lengthening of the EDL changed forces exerted on the tibia and forces exerted by its antagonistic muscle group. Apart from its extramuscular myofascial connections, EDL has no connections to either the tibia or these antagonistic muscles. Proximal EDL lengthening increased distal muscular forces (active PER DeltaF approximately +1.7%), but decreased tibial forces (passive from 0.3 to 0 N; active DeltaF approximately -5%). Therefore, it is concluded that these antagonistic muscles do not act independently, because of myofascial force transmission between them. Such a decrease in tibial force indicates release of pre-strained connections. Distal EDL lengthening had opposite effects (tripling passive force exerted on tibia; active PER force DeltaF approximately -3.6%). It is concluded that the length and relative position of the EDL is a co-determinant of passive and active force exerted at tendons of nearby antagonistic muscle groups. These results necessitate a new view of the locomotor apparatus, which needs to take into account the high interdependence of muscles and muscle fibres as force generators, as well as proximo-distal force differences and serial and parallel distributions of sarcomere lengths that are consequences of such interaction. If this is done properly, the effects of integrating a muscle fibre, muscle or muscle group into higher levels of organisation of the body will be evident.     

Changes in diffraction patterns with length in single muscle fibres at rest

The addition of a simple X-Y sampling circuit to a closed circuit television system (CCTV) permits measurements of a narrow profile of the laser diffraction patterns from single frog muscle fibres at rest. Results confirm that maximum intensity occurs at 2.95–3.00 m, but a positive linear relationship between the dispersions of sarcomeres and sarcomere lengths from 2.7–3.8 m is obtained in four isolated single fibres.     

Filament lengths in frog semitendinosus and tibialis anterior muscle fibres

Summary In frog semitendinosus muscle the descending limb of the length-tension curve is shifted rightward relative to that of tibialis anterior. Both the plateau right corner and the zero-force intercept are equally shifted. To investigate the reason for this shift, we compared filament lengths in the two muscles. Single fibres were mechanically skinned, stretched to reveal filaments clearly, incubated in a solution containing one of several antibodies to enhance filament visualization, and examined by electron microscopy. We found no differences of filament length. Thick filament lengths were 1.62 and 1.61 m, respectively. I-segment lengths were measured by two methods. With the first, filament length was the same for both muscles, 1.95 or 1.98 m, depending on the value taken for the troponin repeat; with the second it was 1.92 and 1.94 m, respectively, for the two muscles. These differences are insignificant. Thus, the reported differences of shape of the length-tension curve are not explainable in terms of differences of filament length.     

Imaging of muscle activity-induced morphometric changes in fibril network of myofascia by two-photon microscopy

Myofascia, deep fascia enveloping skeletal muscles, consists of abundant collagen and elastin fibres that play a key role in the transmission of muscular forces. However, understanding of biomechanical dynamics in myofascia remains very limited due to less quantitative and relevant approaches for in vivo examination. The purpose of this study was to evaluate the myofascial fibril structure by means of a quantitative approach using two-photon microscopy (TPM) imaging in combination with intravital staining of Evans blue dye (EBD), a far-red fluorescence dye, which potentially labels elastin. With focus on myofascia of the tibial anterior (TA) muscle, the fibril structure intravitally stained with EBD was observed at the depth level of collagen fibrous membrane above the muscle belly. The EBD-labelled fibril structure and orientation in myofascia indicated biomechanical responses to muscle activity and ageing. The orientation histograms of EBD-labelled fibrils were significantly modified depending upon the intensity of muscle activity and ageing. Moreover, the density of EBD-labelled fibrils in myofascia decreased with habitual exercise but increased with muscle immobilization or ageing. In particular, the diameter of EBD-labelled fibrils in aged mice was significantly higher. The orientation histograms of EBD-labelled fibrils after habitual exercise, muscle immobilization and ageing showed significant differences compared to control. Indeed, the histograms in bilateral TA myofascia of exercise mice made simple waveforms without multiple sharp peaks, whilst muscular immobilization or ageing significantly shifted a histogram with sustaining multiple sharp peaks. Therefore, the dynamics of fibre network with EBD fluorescence in response to the biomechanical environment possibly indicate functional tissue adaptation in myofascia. Furthermore, on the basis of the knowledge that neutrophil recruitment occurs locally in working muscles, we suggested the unique reconstruction mechanism involving neutrophilic elastase in the myofascial fibril structure. In addition to the elastolytic susceptibility of EBD-labelled fibrils, distinct immunoreactivities and activities of neutrophil elastase in the myofascia were observed after electric pulse stimulation-induced muscle contraction for 15 min. Our findings of EBD-labelled fibril dynamics in myofascia through quantitative approach using TPM imaging and intravital fluorescence labelling potentially brings new insights to examine muscle physiology and pathology.     

An estimate of the mean length of collagen fibrils in rat tail-tendon as a function of age

A theoretical expression has been derived for the mean collagen fibril length in tendon based on the assumption that collagen fibrils originate in cell surface invaginations and terminate either at some remote cell surface or another collagen fibril bundle. The expression thus determined requires knowledge of the effective lengths of the fibrocytes (or fibrocyte assemblies) and the cellular content of the tendon. Both of these parameters have been measured experimentally as a function of age for rat-tail tendon using a combined light microscope and electron microscope approach. The results obtained for immature tendon suggest that the mean collagen fibril length is at least equal to the critical length required to maintain the appropriate tensile properties. In the most mature tissue studied, however, the mean-collagen fibril length is in excess of 100 times the critical length.     

A comparison of isometric force,maximum power and isometric heat rate as a function of sarcomere length in mouse skeletal muscle

The isometric force, maximum power and isometric heat rate have been measured at different sarcomere lengths (SL) between 1.40 and 3.63 m in two types of mouse muscle, soleus and omohyoideus, at 25°C. The SL force relationship is different in the two muscles. At a SL above optimum filament overlap, 2.44 m in omohyoideus muscles, maximum power declined while isometric force remained high. In soleus muscles this occurred above a SL of 2.33 m. In parallel experiments, the isometric heat rate declined linearly with increasing SL above 2.33 m in soleus muscles, while isometric force remained closer to its maximum. At short SL, between 2.33–1.75 m in soleus and 2.44–2.15 m in omohyoideus, maximum power remained at or near its maximum value as did heat rate (soleus) while isometric force fell. In both muscles at SL greater than optimum for force development maximum power output (unlike force) is proportional to filament overlap. The variation in heat rate over this SL range can be described as the sum of a constant rate and a rate proportional to filament overlap. These observations are compatible with the idea that maximum power and heat rate are less affected by non-uniformities in SL than is force.     

Skeletal muscle oedema and muscle fibre necrosis during septic shock. Observations with a porcine septic shock model

In domestic pigs, intermitted application of Escherichia coli-endotoxin was used to create an animal model for a prolonged hypo- and hyperdynamic septic shock-like state and to investigate mechanisms of multiple organ failure. Here, we describe the changes in skeletal muscle after 18 h (2 animals) and 48 h (6 animals) of septic shock. Two pigs for each observation period that received physiologic saline solutions instead of endotoxin served as controls. The earliest lesions were endothelial cell damage with endomysial oedema and swelling of mitochondria in muscle fibres. With increasing degree of endothelial cell damage, pericytes showed degenerative changes with cytoplasmic fragmentation and karyolysis. After 48 h of shock, endomysial oedema was increased with fibrinogen present. Muscle fibre diameters were increased and swollen mitochondria and segmental necrosis of muscle fibres were frequently observed. However, phagocytic reaction or regenerative changes were not detected. In this respect, skeletal muscle lesions in septic shock differ from ischemic damage, which is characterized by early phagocytosis. Tumour necrosis factor alpha (TNF) was increased greatly and significantly in the serum of the pigs that received endotoxin. The lesions described may be the result of both direct damage to muscle fibres by the endotoxin and/or the increased levels of TNF and indirect damage because of the increased diffusion distance, due to the endomysial oedema. The loss of blood proteins into the endomysium may also play a role in generating hypoproteinemia in patients with septic shock.     

Connectin,giant elastic protein,in giant sarcomeres of crayfish claw muscle

Summary In the giant sarcomeres (sarcomere length, 10 m at rest) of crayfish claw muscle, 3000 kDa connectin-like protein but not projectin (mini-titin) appears to be responsible for passive tension generation. Proteolysis of crayfish connectin in skinned fibres was parallel with disappearance of resting tension. Immunofluorescence observations using the antiserum to crayfish connectin showed that crayfish connectin linked the A band to the Z line ina giant sarcomere. It appears that crayfish connectin exerts a centering force on the A band in a sarcomere. Very thin filaments in the I band were visualized after the actin filaments had been removed by the treatment with plasma gelsolin. Crayfish connectin was partially purified and its rotary shadowed image was a very long filament. Projectin was localized on the A band of crayfish giant sarcomeres and remained unmoved during stretch or contraction. However, on dissolution of myosin filaments, projectin moved to the Z line together with crayfish connectin. It seems that projectin binds to connectin on the myosin filament. In regular size of sarcomeres (sarcomere lengths, 3–4 m at rest) of crayfish stretcher muscle, projectin linked the A band to the Z line, as in insect flight muscle.