Skeletal muscle HSP expression in response to immobilization and remobilization.

Heat shock proteins play an important regulatory role in the cellular defence. Oxidative stress is one of the factors inducing heat shock protein expression. This study tested the effects of 4 weeks of immobilization and subsequent remobilization on heat shock protein expression and oxidative stress in the lateral gastrocnemius and plantaris muscles of the rat. Active mobilization or free mobilization protocols were used for remobilization. In active mobilization, strenuous uphill treadmill running, twice a day, was started immediately after the immobilization and lasted for six days. Rats in the free mobilization group moved freely in their cages immediately after the immobilization. Expression of heat shock proteins was upregulated during the recovery from immobilization, especially in the lateral gastrocnemius muscle in the active mobilization group. However, markers of oxidative stress, such as protein carbonyls and 4-hydroxynonenal protein adducts, or activities of the antioxidant enzymes glutathione peroxidase and glutathione reductase, did not change after the immobilization and subsequent recovery. In summary, following immobilization, both intensive and spontaneous exercise upregulated the heat shock protein expressions in the lateral gastrocnemius muscle and partly in the plantaris muscle, which may contribute to the recovery from immobilization atrophy.

Recovery from immobilisation: responses of fast-twitch muscle fibres to spontaneous and intensive exercise in rat calf muscles.

Four weeks of immobilisation with two types of re-mobilisation programmes (intensive concentric treadmill exercising during 6 days, and free exercising, and immobilisation without any re-mobilisation period were studied to clarify possible exercise-induced calf muscle damage especially in fast-twitch fibres used in running compared to non-immobilised rats housing freely in their cages. As markers of muscle injury, conventional histology, beta-glucuronidase (beta-GU) activity and fetal myosin heavy chain expression (MHC-d) were assessed on Days 0, 1, 3, 6 and 14 after the cast removal. Only minor focal hypercontraction, ruptures and necrosis of myofibrils, and weak inflammatory cell reactions were found in all samples examined, except in the controls. No MHC-d positive cells were found indicating absence of active regeneration after immobilisation or re-mobilisation. Minor increase in beta-GU activity was observed in all three muscles studied, but statistically significant increase was observed only in the samples of the free exercising group on Day 14 after the cast removal. To conclude, intensive concentric treadmill exercise for 6 days did not cause significantly more muscle damage than did free exercising re-mobilisation.

Spiral cracks in drying precipitates.

We investigate the formation of spiral crack patterns during the desiccation of thin layers of precipitates in contact with a substrate. This symmetry-breaking fracturing mode is found to arise naturally not from torsion forces but from a propagating stress front induced by the foldup of the fragments. We model their formation mechanism using a coarse-grain model for fragmentation and successfully reproduce the spiral cracks. Fittings of experimental and simulation data show that the spirals are logarithmic. Theoretical aspects of the logarithmic spirals are discussed. In particular we show that this occurs generally when the crack speed is proportional to the propagating speed of stress front.

Effects of remobilization on rat femur are dose-dependent.

Seventy 9 to 11-week-old Sprague-Dawley male rats were divided into seven groups [baseline, 3-week, 11-week control groups; a group with a left limb immobilization for 3 weeks; and three groups with a similar immobilization and subsequent 8-week free (FR), low-intensity running (LR) or high-intensity running (HR) remobilization] to determine the site-specific effects of decreased mechanical loading and subsequent increased activity on rat femur. Bone mineral content of the proximal femur (PBMC), femoral midshaft (SBMC) and distal femur (DBMC) and the histomorphometry of the distal femur were used as outcome variables. The 3-week immobilization period resulted in significant bone loss in the proximal and distal ends of the immobilized left limb, the deficit being -5.9% in PBMC and -14.7% in DBMC. Immobilization also led to marked microarchitectural changes in the distal femur, the left-side deficit in trabecular bone volume (BV/TV) being -23.9%. After remobilization, there was a clear indication for dose-dependent response; i.e., immobilization-induced left-to-right side differences in BV/TV persisted in the FR animals (-23.8%), while these left limb deficits were significantly reduced in the LR group (-15.0%) and virtually absent in the HR group (-3.1%). The left limb deficit in PBMC was still significant in all groups after the 8-week period of remobilization [-9.6% in the FR group; -13.4% in the LR group; and -7.2% in the HR group]. The left-limb deficit in SBMC was significant in the HR11 group only (-7.2%), and, contrary to histomorphometric data, virtually absent in DBMC in all remobilization groups. As compared with the age-matched control data, the weight-adjusted BMCs of both limbs of the LR and HR groups were comparable or even higher (right limbs) than those of the controls. In conclusion, this study indicates that remobilization-induced bone recovery depends on the intensity of the remobilization so that during the 8-week period of remobilization, high-intensity running results in better recovery than low-intensity running, both of which are more efficient than free-cage activity only. Immobilization-induced changes in rat femur are also restored in a site-specific fashion, the most trabecular distal region of the femur showing more complete recovery than the more cortical proximal and midshaft regions.

Achilles tendon injuries.

The Achilles tendon is the strongest tendon in the human body. Because most Achilles tendon injuries take place in sports and there has been an common upsurge in sporting activities, the number and incidence of the Achilles tendon overuse injuries and complete ruptures have increased in the industrialized countries during the last decades. The most common clinical diagnosis of Achilles overuse injuries is tendinopathy, which is characterized by a combination of pain and swelling in the Achilles tendon accompanied by impaired ability to perform strenuous activities. Most patients with Achilles tendon injury respond favorably to conservative treatment and only those who fail to respond to carefully followed nonoperative treatment should undergo surgery for repair. A complete rupture of the Achilles tendon usually occurs in sports that require jumping, running, and quick turns. Although histopathologic studies have shown that ruptured Achilles tendons include clear degenerative changes before the rupture, many of the Achilles tendon ruptures occur suddenly without any preceding signs or symptoms. Neither conservative nor operative treatment is a treatment of choice for the ruptured Achilles tendon. It is generally accepted that surgery should be performed on ruptured Achilles tendons in young, physically active patients and in those patients for whom the diagnosis or the treatment of the rupture has been delayed, whereas the results of conservative treatment are an acceptable outcome in older patients with sedentary lifestyles. Many important issues still remain unanswered concerning the cause, pathogenesis, diagnosis, and management of the Achilles tendon disorders. Only when these issues have been solved by well-controlled studies can tailored treatment protocols be created.

Tenascin-C in the pathobiology and healing process of musculoskeletal tissue injury.

Tenascin-C (TN-C) is a hexabrachion-shaped extracellular matrix (ECM) protein which has very restricted expression in normal musculoskeletal tissues, but is expressed in large quantities in these tissues during embryogenesis as well as during regenerative and healing processes. TN-C is an elastic protein which has a number of binding sites for other extracellular matrix proteins as well as for cell membrane adhesion receptors. In addition, it can be stretched to several times its resting length, the ability of which is attributed to the stretch-induced unfolding of its fibronectin type III domains. In the musculoskeletal tissues, TN-C is present in the regions where high mechanical forces are transmitted from one tissue component to another, such as the myotendinous and osteotendinous junctions. Not surprisingly, it was recently presented that the expression of TN-C in the musculoskeletal tissues is regulated by the mechanical strain applied to their cells. Thus, taking into account the flexible structure of the TN-C and its site-specific expression pattern at sites exposed to heavy mechanical loads, TN-C is likely to play an important role in providing elasticity to the musculoskeletal tissues. This feature has a special significance in the degenerative and regenerative processes where the normal biomechanical environment of the musculoskeletal tissue is temporarily interrupted by injury. The rapidly increasing understanding of the structure and function of the ECM protein TN-C may well bring important insights into the clinical treatment of sports injuries in the future.

[Microscopic study of dental calculus in cadavers from the 18th-19th centuries]. / 18-19. századi múmiák fogköveinek mikroszkópos vizsgálata.

The authors studied the dental calculus of 20 mummies with ligth microscopy, polarized ligth microscopy and scanning electron microscopy. Gram positive bacteria could be detected in all preparates, while Gram negative bacteria in 12 and fungi only in 3 dental calculus was visible. Animal food remains within five, and plant remains in all dental calculus were identified. Anorgic element and cell debris were seen in all preparates.

Mechanical loading regulates tenascin-C expression in the osteotendinous junction.

Elastic extracellular matrix protein tenascin-C (TN) has very restricted expression in normal tissues, but is expressed in large quantities during embryogenesis and hyperplastic processes. To examine the importance of mechanical stress on the regulation of TN expression in vivo, the effects of various mechanical loading states (immobilization and three forms of subsequent remobilization) on the expression of TN were studied immunohistochemically at the bone-tendon attachment of the rat quadriceps muscle. This osteotendinous junction (OTJ) was selected as study site, since it receives its mechanical stimuli only from muscle contracting activity, which is easy to block by cast immobilization. TN was expressed abundantly in the normal OTJ. Following the removal of the mechanical stress from the junction by cast-immobilization of three weeks, the immunoreactivity of TN was almost completely absent. Normal mechanical stress in the form of free remobilization of eight weeks (free cage activity) resulted in a slight increase in TN expression, but could not restore the expression of TN to the level of the healthy contralateral leg. After the application of the increased mechanical stress (intensified remobilization of the eight weeks by low- or high-intensity treadmill running), the distribution and immunoreactivity of TN reached the level of the healthy contralateral limb in the low-intensity running group or even exceeded that in the high-intensity running group. High TN expression was seen around the chondrocytes and fibroblasts of the OTJ as well as around the collagen fibers of the tendon belly. We conclusively show that mechanical strain regulates the expression of TN in vivo, and propose that mechanical stress is a major regulator of TN expression in fibroblasts and chondrocytes. This may be an important aspect of the regulation of TN expression during embryogenesis, tendon degeneration, wound healing, bone formation, and in the other normal or regenerative morphogenetic processes TN is postulated to take part in.

Effects of immobilization and subsequent low- and high-intensity exercise on morphology of rat calf muscles.

After a cast immobilization of 3 weeks, the effects of 4-week remobilization by free cage activity or treadmill running on the morphology of the rat soleus and gastrocnemius muscles were studied. The studied morphometric parameters were: percentage volume of intramuscular connective tissue, capillary density, muscle fiber size, number of fibers with a pathological structural alteration, and fiber type distribution. In both muscles, immobilization of 3 weeks produced a significant increase in the connective tissue volume and number of fibers with pathological alterations, with a similar decrease in the capillary number and fiber size. At the same time, the relative amount of type I fibers decreased and type IIA fibers increased. Free remobilization and especially intensified remobilization by treadmill running significantly restored these values towards controls. These findings indicate that in rat soleus and gastrocnemius muscles immobilization-induced accumulation of intramuscular connective tissue, capillary loss, reduction in fiber size, accumulation of fibers with pathological structural alterations, and changes in fiber type distribution are to a great extent reversible phenomena, especially if remobilization is intensified by physical training. In clinical practice, this suggests that in patients with musculoskeletal injuries the postimmobilization rehabilitation should be early and effective.

Blood flow in rat gastrocnemius muscle and Achilles tendon after Achilles tenotomy.

The effect of Achilles tenotomy on resting blood flow of rat gastrocnemius muscle and Achilles tendon was studied by radioactive microspheres. Tenotomy produced an immediate, marked decrease in both intramuscular and intratendinous blood flow and it remained significantly lowered at both sites till the end of the observation period, i.e., day 18 after tenotomy. The decrease in the resting blood flow was more rapid and pronounced in the Achilles tendon than in the gastrocnemius muscle. Although the blood flow of the Achilles tendon started to recover after the 4th postoperative day, it was still 33% (statistically not significant) lower than that in the controls 18 days after tenotomy. In the gastrocnemius muscle, the 18-day deficit was 38% (p < 0.001), respectively. The results indicate that after division of a rat Achilles tendon the resting blood flow to the gastrocnemius muscle and Achilles tendon is not adequately restored, remaining at a significantly lowered level even 18 days after tenotomy.

Free mobilization and low- to high-intensity exercise in immobilization-induced muscle atrophy.

After 3 wk of immobilization, the effects of free cage activity and low- and high-intensity treadmill running (8 wk) on the morphology and histochemistry of the soleus and gastrocnemius muscles in male Sprague-Dawley rats were investigated. In both muscles, immobilization produced a significant (P < 0.001) increase in the mean percent area of intramuscular connective tissue (soleus: 18.9% in immobilized left hindlimb vs. 3.6% in nonimmobilized right hindlimb) and in the relative number of muscle fibers with pathological alterations (soleus: 66% in immobilized hindlimb vs. 6% in control), with a simultaneous significant (P < 0.001) decrease in the intramuscular capillary density (soleus: mean capillary density in the immobilized hindlimb only 63% of that in the nonimmobilized hindlimb) and muscle fiber size (soleus type I fibers: mean fiber size in the immobilized hindlimb only 69% of that in the nonimmobilized hindlimb). Many of these changes could not be corrected by free remobilization, whereas low- and high-intensity treadmill running clearly restored the changes toward control levels, the effect being most complete in the high-intensity running group. Collectively, these findings indicate that immobilization-induced pathological structural and histochemical alterations in rat calf muscles are, to a great extent, reversible phenomena if remobilization is intensified by physical training. In this respect, high-intensity exercise seems more beneficial than low-intensity exercise.

Location and distribution of non-collagenous matrix proteins in musculoskeletal tissues of rat.

The study assessed immunohistochemically the location and distribution of various non-collagenous matrix proteins (fibronectin, laminin, tenascin-C, osteocalcin, thrombospondin-1, vitronectin and undulin) in musculoskeletal tissues of rat. Fibronectin and thrombospondin-1 were found to be ubiquitous in the studied tissues. High immunoreactivity of these proteins was found in the extracellular matrix of the anatomical sites where firm bindings are needed, i.e. between muscle fibres and fibre bundles, between the collagen fibres of a tendon and at myotendinous junctions, osteotendinous junctions and articular cartilage. Tenascin-C was found in the extracellular matrix of regions where especially high forces are transmitted from one tissue component to the other, such as myotendinous junctions and osteotendinous junctions. Laminin was demonstrated in the basement membranes of the muscle cells and capillaries of the muscle-tendon units. Osteocalcin immunoreactivity concentrated in the extracellular matrix of areas of newly formed bone tissue, i.e. in the subperiosteal and subchondral regions, osteoid tissue and mineralized fibrocartilage zone of the osteotendinous junction. Mild vitronectin activity could be seen in the extracellular matrix of the osteotendinous and myotendinous junctions, and high activity around the bone marrow cells. Undulin could be demonstrated in the extracellular matrix (i.e. on the collagen fibres) of the tendon and epimysium only. However, it was co-distributed with fibronectin and tenascin-C. Together, these findings on the normal location and distribution of these non-collagenous proteins in the musculoskeletal tissues help to form the basis of knowledge against which the location and distribution of the these proteins in various pathological processes could be compared.

Effects of training, immobilization and remobilization on tendons.

Since a tendon is a living tissue, it is not a surprise that tendon shows the capacity to adapt its structure and mechanical properties to the functional demands of the entire muscle-tendon unit. However, compared with muscle, the experimental knowledge of the effects of strength or endurance-type training on tendon tissue is scarce and clinical human experiments are completely lacking (1). Research should, however, be able to improve the true understanding of the biomechanical, functional, morphological and biochemical changes that occur in tendons due to training and physical activity, since understanding of the basic physiology of a tissue is the key to understanding its pathological processes (1, 2). Compared with muscle tissue, the metabolic turnover of tendon tissue is many times slower due to poorer vascularity and circulation (1, 3). The adaptive responses of tendons to training are therefore also slower than those in muscles, but they may finally be considerable if the time frame is long enough (3, 4).

Histopathological findings in chronic tendon disorders.

Tendon injuries and other tendon disorders represent a common diagnostic and therapeutic challenge in sports medicine, resulting in chronic and long-lasting problems. Tissue degeneration is a common finding in many sports-related tendon complaints. In the great majority of spontaneous tendon ruptures, chronic degenerative changes are seen at the rupture site of the tendon (1). Systemic diseases and diseases specifically deteriorating the normal structure of the tendon (i.e. foreign bodies, and metabolic, inherited and infectious tendon diseases) are only rarely the cause of tendon pathology. Inherited diseases, such as various hereditary diseases with disturbed collagen metabolism and characteristic pathological structural alterations (Ehlers-Danlos syndrome, Marfani syndrome, homocystinuria (ochronosis)), represent approximately 1% of the causes of chronic tendon complaints (2), whereas foreign bodies are somewhat more common and are found in less than 10% of all chronic tendon problems (1). Rheumatoid arthritis and sarcoidosis are typical systemic diseases that cause chronic inflammation in tendon and peritendinous tissues. Altogether, these 'specific' disorders represented less than 2% of the pathological alterations found in the histological analysis of more than 1000 spontaneously ruptured tendons (1, 3, 4). In this material, degenerative changes were seen in a great majority of the tendons, indicating that a spontaneous tendon rupture is a typical clinical end-state manifestation of a degenerative process in the tendon tissue. The role of overuse in the pathogenesis of chronic tendon injuries and disorders is not completely understood. It has been speculated that when tendon is overused it becomes fatigued and loses its basal reparative ability, the repetitive microtraumatic processes thus overwhelming the ability of the tendon cells to repair the fiber damage. The intensive repetitive activity, which often is eccentric by nature, may lead to cumulative microtrauma which further weakens the collagen cross-linking, non-collagenous matrix, and vascular elements of the tendon. Overuse has also been speculated to cause chronic tendon problems, by disturbing the micro- and macrovasculature of the tendon and resulting in insufficiency in the local blood circulation. Decreased blood flow simultaneous with an increased activity may result in local tissue hypoxia, impaired nutrition and energy metabolism, and together these factors are likely to play an important role in the sequence of events leading to tendon degeneration (4). A sedentary lifestyle has been proposed as a main reason for poor basal circulation of the tendon, and presumably is at least partly responsible for the high number of tendon problems in people with a sedentary lifestyle who occasionally take part in high physical activity sports events.

Histopathological findings in spontaneous tendon ruptures.

A spontaneous rupture of a tendon may be defined as a rupture that occurs during movement and activity, that should not and usually does not damage the involved musculotendinous units (1). Spontaneous tendon ruptures were uncommon before the 1950s. Böhler found only 25 Achilles tendon ruptures in Wien between 1925 and 1948 (2). Mösender & Klatnek treated 20 Achilles tendon ruptures between 1953 and 1956, but 105 ruptures between 1964 and 1967 (3). Lawrence et al. found only 31 Achilles tendon ruptures in Boston during a period of 55 years (1900-1954) (4). During the recent decades tendon ruptures have, however, become relatively common in developed countries, especially in Europe and North America. A high incidence of tendon ruptures has been reported in Austria, Denmark, Finland, Germany. Hungary, Sweden, Switzerland and the USA; somewhat lower incidences have been reported in Canada, France, Great Britain and Spain. On the other hand, Greece, Japan, the Netherlands and Portugal have reported a clearly lower incidence. Interestingly, Achilles tendon ruptures are a rarity in developing countries, especially in Africa and East-Asia (5). In many developed countries, the increases in the rupture incidence have been dramatic. In the National Institute of Traumatology in Budapest, Hungary, the number of patients with an Achilles tendon rupture increased 285% in men and 500% in women between two successive 7-year periods, 1972-1978 and 1979-1985 (5).

Effects of immobilization, three forms of remobilization, and subsequent deconditioning on bone mineral content and density in rat femora.

Disuse is associated with bone loss, which may not be recoverable. It is not known whether intensified remobilization is beneficial in restoring disuse-related bone loss nor if any such benefit would depend upon continuing mobilization for its maintenance. After an immobilization period of 3 weeks, the effects of free remobilization (11 weeks), and low-and high-intensity treadmill running (11 weeks) with and without subsequent deconditioning (18 weeks) on the bone mineral content (BMC) and density (BMD) of the hindlimb femora of Sprague-Dawley rats (n = 98) were studied using a dual-energy X-ray absorptiometric (DXA) scanner. Our hypothesis was that intensified remobilization is beneficial in restoring the BMC and BMD from disuse to normal while subsequent deconditioning is deleterious to these parameters. Immobilization for 3 weeks produced a significant BMC and BMD loss in the immobilized left femur (range -4.4 to -12.8%; p < 0.05-0.001). In the groups with free remobilization (free cage activity), the body weight-adjusted BMCs and BMDs always remained below those in the controls (range -2.3 to -12.1%; p values ranging from NS to < 0.01). Both low- and high-intensity running restored BMC and BMD in the immobilized limb, the effect being better in the latter group. In both of these groups, the values of the immobilized left limbs and those of the free right limbs exclusively exceeded the corresponding values of the age-matched control rats (left limb values 3.0-21.1% higher with p values ranging from NS to < 0.01; right limb values 7.9-21.4% higher with p < 0.05-0.01). However, after the deconditioning period of 18 weeks, the above described beneficial effects of low- and high-intensity running were lost, the left and right limb BMC and BMD values being lower than those in the age-matched controls (range -3.8 to -8.7%; p values ranging from NS to < 0.05). In conclusion, this study clearly indicates the need for greater than normal activity to restore the BMC and BMD after disuse to normal levels. However, the benefits of intensified remobilization are lost if the activity is terminated, and therefore, after immobilization and disuse, bone loading activities should be continued, perhaps indefinitely.