Journal of Orthopaedic Research 8541-547 Raven Press, Ltd., New York 0 1990 Orthopaedic Research Society
Chronic Achilles Paratenonitis with Tendinosis: An Experimental Model in the Rabbit Clas Backman, *Lennart Boquist, Jan FridCn, 'fRonny Lorentzon, and tGoran Toolanen Department of Hand and Plastic Surgery, *Department of Pathology, and TDepartment of Orthopedic Surgery, University of UmeH, UmeH, Sweden
Summary: An experimental model for inducing chronic Achilles paratenonitis with tendinosis in the rabbit is presented. Thirteen rabbits were exercised in a kicking machine producing passive flexions and extensions of the ankle joint. Active contractions of the triceps surae muscles were induced by electric stimulation via surface electrodes. The animals were exercised for 5 to 6 weeks, with a rate of 150 flexions and extensions per minute for 2 h, three times a week. Light microscopic examination showed degenerative changes of the tendon, and increased number of capillaries, infiltrates of inflammatory cells, edema, and fibrosis in the paratenon. We conclude that chronic Achilles paratenonitis with tendinosis can be experimentally induced in a standardized manner in rabbits. Key Words: Experimental model-ParatenonitisPeritendinitis-Tendinitis-Tendinosis-Histopathology.
Achilles tendon paratenonitis is a common cause of complaint in long-distance runners (3,5,9,10,18). Despite the high incidence of Achilles paratenonitis, with or without tendinosis, the associated histopathological alterations are sparsely described in the literature (2,4). In the literature, the terms peritendinitis, paratenonitis, and tendinitis have been used to describe inflammatory alterations in the paratenon. We have used paratenonitis to describe the alterations in the loose areolar tissue surrounding the tendon with its epitenon, and tendinosis to describe the degenerative alterations in the tendon (15). Experimental models for producing acute paratenonitis have previously been presented by Obolenskaja and Goljanitski (14) and Rais (16). In the study of Obolenskaja and Goljanitski (14), the hind legs of nonanesthetized rabbits were subjected to passive movements for periods ranging from 1 to 6
h, at a rate of 300 movements per minute. Rais (16) exercised rabbits for 0.5 to 6 h with a frequency of 150 movements per minute and used electrical stimulation of the sciatic nerve to accomplish an active contraction. The latter author demonstrated slight edema in muscle, paratenon, and mesotenon after 0.5 h of exercise. The degree of these changes correlated to the duration of the exercise. Perivascular accumulations of polymorphonuclear leukocytes and an increased number of fibroblasts in the paratenon and mesotenon were noted on histopathological examination of specimens from rabbits killed 4 days after an exercise period of 6 h. In this study, an experimental model in the rabbit was designed and tested in an attempt to standardize the conditions related to the development of chronic Achilles paratenonitis with tendinosis. MATERIAL AND METHODS
The experiment was carried out on 13 New Zealand white rabbits of both sexes aged from 6 to 9 months. The animals were caged all of their lives.
Address correspondence and reprint requests to Dr. C. Backman, Department of Hand Surgery, University Hospital of Umel, S-901 85 Umel, Sweden.
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Their weights ranged from 2.5 to 4 kg. Before and after exercise, the animals were kept in ordinary cages allowing good freedom of movement and had free access to food and water. Throughout the experiment, the rabbits were kept under anesthesia, induced by intramuscular injection of fentanylfluanison (0.3 ml/kg) and diazepam (2 mg/kg). This was followed by another injection of fentanylfluanison (0.15 ml/kg) 30 min later. A kicking machine (Fig. 1) was used for passive flexions and extensions of the ankle joint. The movements were produced by a pneumatic piston, where the range of movement could be altered. The movement was set to 9.5 cm, giving a range of motion in the ankle of 55-65", of which 20-25" was dorsiflexion and 3540" was plantar flexion. In order to attain this angular movement, a band was tied around the thigh of the rabbit to restrict the motion of the hip and knee. The piston was attached to the right foot. The left leg served as control and was at rest throughout the experiment. The machine was constructed for the simultaneous exercising of three rabbits. An active contraction was induced during the plantar flexion phase by electrical stimulation via surface electrodes (Pediatric electrodes 40 426A, Hewlett Packard, Andover, MA, U.S.A.) placed 2 cm apart over the triceps surae muscles. This stimulation was synchronized with the plantar flexion movement of the piston by a microswitch, which
5 A
7
1
trigged the stimulator unit (Disa stimulator Type 14E 10, Disa Elekronik A / S , Herlev, Denmark). The stimulator was set to generate an impulse train of 92 ms (impulse duration of 0.2 ms at 18 ms intervals), giving rise to six pulses per train. The pattern of the stimulus was set after pilot experiments. The muscle contraction was triggered 130 ms before the piston was reversed. The time between muscle stimulation and peak tension (6) makes the triceps surae muscles contract during the prestrech and the initiation of the passive dorsiflexion movement. The electrical stimulus was controlled by oscillometer. We used the same setup in all experiments, except for the amplitude, which ranged between 35 and 50 V, depending on the individual contraction response. Throughout the experiment, the contraction response was controlled and if necessary the amplitude was adjusted during the exercise in order to achieve a constant contraction. In a pilot study, five rabbits were exercised for 1 h three times a week for 1 to 6 weeks and the tendon was palpated regularly. The frequency of the flexion and extension movements was set to 150 per minute. Following 6 weeks of exercise, inflammatory alterations in the paratenon was the major histopathological finding. Lymphocytes predominated among the inflammatory cells. The signs of degeneration in the tendon, with variation in staining affinity and varying thickness of the fibrillar components, were present but only to a minor extent. Based on clinical and histopathological findings, 2 h of exercise three times a week for 5 to 6 weeks was used for this study. After the animals were sacrificed (the day after the last exercise), the Achilles tendons from both legs were excised and placed in formalin. The specimen was kept in formalin for 2-3 days. After dehydration in alcohol, the tendon was embedded in paraffin and longitudinally cut (section thickness of 5-6 Fm). The sections were stained with hematoxylin-eosin.
RESULTS
FIG. 1. (A) Rabbits positioned in kicking apparatus. (1) DlSA stimulator unit. (2) Switch for individual adjustment of amplitude. (3) Pneumatic piston. (4) Band restricting the movement of hip and knee. (B) Closeup view of hindfoot and pneumatic piston. Microswitches (5) regulate the motion length of the piston and switch (6) triggers the DlSA stimulator unit. Surface electrodes placed on the triceps surae muscles (7).
J Orthop Res, Vol. 8, No. 4, 1990
Throughout the experimental period, the animals were in generally good condition. After 4 weeks of exercise, palpation of the exercised leg indicated that all animals had irregular thickening over the Achilles tendon. At this time, most animals demonstrated palpable nodules 0.5 to 1 cm above the insertion into the calcaneus. Light microscopic examination disclosed a normal histological appearance of tendon and paratenon in the control legs (Figs. 2A and 3A).
EXPERIMENTAL CHRONIC ACHILLES PARA TENONITIS The tendons of the exercised legs exhibited degenerative changes of varying severity and distribution. Most often, these changes were localized to the central portion of the tendons (Fig. 2B). The staining affinity varied in different portions of the tendons, and the fibrillar components showed varying thickness (Fig. 2C, 2D). The nuclei of the ten-
543
dons exhibited slightly varying size, contour, and staining affinity. Fibrillation was occasionally encountered in the tendons (Fig. 3B). The paratenon of the exercised legs was thickened (Figs. 4A-D and 5A) with the occurrence of an increased number of fibroblasts (Figs. 3B and 4AD). The number of capillaries was also increased
A.B
FIG. 2. (A) Low-power micrograph demonstrating a normal appearance of tendon and thin paratenon (to the right). Control leg, 6 weeks. Hematoxylin-eosin, x25. (6)Low-power micrograph showing degenerative changes with varying staining affinity in the tendon, and thickened paratenon (to the right). Exercised leg, 6 weeks. Hematoxylin-eosin, ~ 3 7 (C) . Micrograph showing degenerative tendon. Exercised leg, 5 weeks. Hematoxylin-eosin, ~ 1 5 0(D) . Higher magnification demonstrating degenerative alterations in tendon with varying staining affinity. Exercised leg, 6 weeks. Hematoxylin-eosin, ~237.
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FIG. 3. (A) Normal tendon and paratenon. Control leg, 6 weeks. Hernatoxylin-eosin, x375. (B) Micrograph demonstrating fibrillation in tendon, dilated capillary (C) containing erythrocytes, and increased number of firoblasts (F)in paratenon. Exercised leg, 5 weeks. Hematoxylin-eosin, x375.
(Figs. 4C and 5A), and these vessels were frequently dilated (Fig. 5A) and filled with erythrocytes (Fig 3B). Additional changes in the paratenon of the exercised legs were edema (Figs. 5A, 5B) and infiltration of inflammatory cells (Figs. 5B-D). Lymphocytes predominated in the inflammatory cell idiltrates, but plasma cells and granulocytes were also encountered. A varying number of eosinophils could be observed, in addition to neutrophils. Histiocytes were also found. A distinct perivascular distribution of inflammatory cells was a common finding (Figs. 5D). Table 1 shows a semiquantitative assessment of the histopathological changes observed in each experimental animal. DISCUSSION
This study demonstrates an animal model for inducing histopathological changes associated with chronic Achilles paratenonitis and paratenonitis with tendinosis, i.e., degenerative alterations of tendon, inflammatory cells, increased number of capillaries, as well as edema and fibrosis in the paratenon (15). Although the extent of histopathological change varied among the experimental animals, all of these animals exhibited alterations both
J Orthop Res, Vol. 8, No. 4, 1990
in tendon and paratenon as demonstrated in a semiquantitative manner in Table 1. Previous experimental studies have focused upon the acute Achilles paratenonitis (14,16). In order, however, to correlate to the prolonged overuse situation in humans, predisposing for tendon ruptures (4,5,11,17), a more extended period of stimulus imposed on the tendon is necessary (2). The choice of this protocol was based on preliminary experiments indicating that 1 h of exposure (three times per week for 6 weeks) to exercise brought about inflammatory changes in the paratenon but merely minor signs of degeneration within the tendon (Backman et al., unpublished data). Also, pilot experiments showed that 4 weeks of exercise gave rise to palpatory changes, i.e., nodules around the tendon and diffuse thickening of the soft tissues that are in concert with the observations reported by Kvist et al. (8). In order to mimic the etiological factors in humans, the rabbit tendons were subjected to simultaneous electrical muscle stimulation and traction of the Achilles tendon. This regimen has previously been shown to be the most successful way of inducing acute inflammatory response in the Achilles paratenon (16). The stimulation protocol, giving impulse trains of 92 ms duration, indicates the necessity of a more extended contraction, as opposed to
EXPERIMENTAL CHRONIC ACHILLES PARA TENONITIS
545
C.D
FIG. 4. (A) Low-power micrograph showing markedly thickened paratenon (P), increased number of capillaries (C) and adipose tissue (A). The tendon is seen to the left. Exercised leg, 6 weeks. Hematoxylin-eosin, ~ 4 3(6) . Higher magnification of thickened paratenon showing numerous fibroblasts (F). Adipose tissue (A) and tendon (T) are also seen. Exercised leg, 6 weeks. Hematoxylin-eosin, x95. (C) Micrograph showing thickened paratenon with increased number of capillaries and fibroblasts. Exercised leg, 6 weeks. Hematoxylin-eosin, x95. (D) Micrograph showing paratenon with increased number of fibroblasts and fibrils. Exercised leg, 5 weeks. Hematoxylin-eosin, x375.
a single twitch. The stimulation during the plantar flexion phase creates a contraction of the myotendinous tissue while in a shortened position and through the initiation of the dorsiflexion movement (6). This is in line with recent studies demonstrating
that during running as well as normal stride, a prestretch of the gastrocnemii occurs prior to foot placement when landing (13). To induce hyperfunction, we applied the same kicking frequency as that implemented to cause
J Orthop Res, Vol. 8, No. 4, 1990
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FIG. 5. (A) Micrograph demonstrating wavy appearance in tendon (T) and markedly thickened paratenon with dilated capillaries (C),edema (E), and fibrosis. Exercised leg, 5 weeks. Hematoxylin-eosin, x60. (B) Higher magnification showing edema (E) and inflammatory cells (I), including lymphocytes and granulocytes, and some fibroblasts. Exercised leg, 5 weeks. Hematoxylineosin, x375. (C) A rich number of inflammatory cells, mainly lymphocytes, but also some plasma cells and granulocytes, is interspersed among connective tissue fibrils and small edematous areas in paratenon. Exercised leg, 5 weeks. Hematoxylineosin, x375. (D)Micrograph demonstrating perivascular infiltration of inflammatory cells in adipose tissue close to a tendon. Exercised leg, 5 weeks. Hematoxylin-eosin, ~ 2 3 7 .
acute paratenonitis (16). Due to the fact that this is a survival experiment, we modified the stimulation technique and employed transcutaneous electrical stimulation of the muscles, as opposed to the indirect sciatic nerve stimulation used by Rais (16). A higher stimulation voltage is necessary to proJ Orthop Res, Vol. 8, No. 4, 1990
duce an active contraction when applying transcutaneous muscle stimulation as compared to nerve stimulation. The transcutaneous stimulation is, however, easily reproduced, noninvasive, and convenient for repeated application. It may be argued that the transcutaneous stimulation per se could
547
EXPERIMENTAL CHRONIC ACHILLES PARATENONITIS TABLE 1. Semiquantitative assessment of histopathological changes in each experimental animal Tendon Animal no. 6 7 8 9 10 11 12 13 14 15
16 17 18
0
=
Paratenon
Staining affinity
Nuclear appearance
Fibrillar appearance
2 1 3 3 1 0 3 2 0 3 2 3 3
1 1 2 1 2 2 2 3 0 3 2 1 2
1 2 3 2 2 1 3 2 1 2 3
1 2 1 3 3 0 2 1 3 2
1
2 3
no change, 1
=
slight change, 2
3 =
Thickness
3
moderate change, and 3
cause an inflammatory reaction in the tendon. There are two major objections to such an assumption. First, the electrodes were located at some distance from the tendon. Second, changes related to electrical stimulation appear to be confined to muscle tissue (1,12). Our findings are identical to those reported in biopsy material from professional runners and joggers with sustained Achilles tendon complaints, admitted for surgery after months or years of nonbeneficia1 conservative treatment (2). The described biopsy material shows paratendineal thickening, edema, proliferation of fibroblasts, formation of new connective tissue, infiltrates of inflammatory cells, and proliferation of small blood vessels in the paratenon (7). The inflammatory changes in paratenon are often combined with areas of focal degeneration within the tendon, indicating paratenonitis with tendinosis (2). The presence and degree of edema and polymorphonuclear leukocytes is a sign of acute peritendinis. This result is explained by the fact that the animals were sacrificed the day after their last exercise bout. In conclusion, this model provides a tool for the further understanding of the mechanisms involved in the development and possibly in the treatment of chronic Achilles paratenonitis with tendinosis. Acknowledgment: This work has been supported by grants from the Research Council of the Swedish Sports Federation and the Medical Faculty, Umei University.
REFERENCES 1. Cabric M, Appell HJ, Resic A: Effects of electrical stimula-
tion of different frequencies on the myonuclei and fiber size in human muscle. Int J Sports Med 8:323-326, 1987
=
Occurrence of fibrosis
Occurrence of edema
1 2 I 3 2 0 2 2 2 2 3 1 2
0 1 0 1 2 3
0 1 0 0
1 2 3
Capillaries
Inflammation
2 2 3 1 3 3 1 2 2 1 2 3 2
3 2 3 2 3 3 2 1 0 1 3 2 2
marked change. 2. Clancy WG, Neidhart D, Brand RL: Achilles tendonitis in runners: A report of five cases. Am J Sports Med 4:46-57, 1976 3. Clement DB, Taunton JE, Smart GW: Achilles tendinitis and peritendinitis: Etiology and treatment. Am J Sports Med 12:179-184, 1984 4. Davidsson L, Salo M: Pathogenesis of subcutaneous tendon ruptures. Acta Chir Scand 135:20%212, 1969 5. Fox JM, Blazina ME, Jobe FW, Kerlan RK, Carter VS, Shields CL, Carlson GJ: Degeneration and rupture of the Achilles tendon. Clin Orthop 107:221-224, 1975 6. Freund HJ: Motor unit and muscle activity in voluntary motor control. Physiol Rev 63:387436, 1983 7. Kvist M, Jozsa L, Jarvinen M, Kvist H: Chronic achilles paratenonitis in athletes: A histological and histochemical study. Pathology 19:1-1 1 , 1987 8. Kvist H, Kvist M: The operative treatment of chronic calcaneal paratenonitis. J Bone Joint Surg [Br] 62:353-357, 1980 9. Leach RE, DiIorio E, Harney RA: Pathologic hindfoot conditions in the athlete. Clin Orthop 177:116-121, 1983 10. Leach RE, James S, Wasilewski S: Achilles tendinitis. Am J Sports Med 9:93-98, 1981 1 1 . Ljungkvist R: Subcutaneous partial rupture of the Achilles tendon. Acta Orthop Scand Suppl 113:1-86, 1968 12. Myrhage R, Hudlicka 0: Capillary growth in chronically stimulated adult skeletal muscle as studied by intravital microscopy and histological methods in rabbits and rats. Microvasc Res 16:73-90, 1978 13. Nilsson J, Thorstensson A, Halbertsma J: Changes in leg movements and muscle activity with speed of locomotion and mode of progression in humans. Acta Physiol Scand 123:457-476, 1985 14. Obolenskaja AJ, Goljanitzki JA: Die serose Tendovaginitis in der Klinik und im Experiment. Dtsch Z Chir 201:388-399, 1927 15. Puddu G, Ippolito E, Postacchini F: A classifcation of achilles tendon disease. Am J Sports Med 4:145-150, 1976 16. Rais 0:Heparin treatment of peritenomyosis (peritendinitis) crepitans acuta. Acta Chir Scand Suppl268: 1-88, 1961 17. Skeoch DU: Spontaneous partial subcutaneous ruptures of the tendo achilles. Am J Sports Med 9:20-22, 1981 18. Snook GA: Achilles tendon tenosynovitis in long distance runners. Med Sci Sporrs 4:155-158, 1972
J Orthop Res, Vol. 8, No. 4, 1990
Chronic Achilles Paratenonitis with Tendinosis: An Experimental Model in the Rabbit Clas Backman, *Lennart Boquist, Jan FridCn, 'fRonny Lorentzon, and tGoran Toolanen Department of Hand and Plastic Surgery, *Department of Pathology, and TDepartment of Orthopedic Surgery, University of UmeH, UmeH, Sweden
Summary: An experimental model for inducing chronic Achilles paratenonitis with tendinosis in the rabbit is presented. Thirteen rabbits were exercised in a kicking machine producing passive flexions and extensions of the ankle joint. Active contractions of the triceps surae muscles were induced by electric stimulation via surface electrodes. The animals were exercised for 5 to 6 weeks, with a rate of 150 flexions and extensions per minute for 2 h, three times a week. Light microscopic examination showed degenerative changes of the tendon, and increased number of capillaries, infiltrates of inflammatory cells, edema, and fibrosis in the paratenon. We conclude that chronic Achilles paratenonitis with tendinosis can be experimentally induced in a standardized manner in rabbits. Key Words: Experimental model-ParatenonitisPeritendinitis-Tendinitis-Tendinosis-Histopathology.
Achilles tendon paratenonitis is a common cause of complaint in long-distance runners (3,5,9,10,18). Despite the high incidence of Achilles paratenonitis, with or without tendinosis, the associated histopathological alterations are sparsely described in the literature (2,4). In the literature, the terms peritendinitis, paratenonitis, and tendinitis have been used to describe inflammatory alterations in the paratenon. We have used paratenonitis to describe the alterations in the loose areolar tissue surrounding the tendon with its epitenon, and tendinosis to describe the degenerative alterations in the tendon (15). Experimental models for producing acute paratenonitis have previously been presented by Obolenskaja and Goljanitski (14) and Rais (16). In the study of Obolenskaja and Goljanitski (14), the hind legs of nonanesthetized rabbits were subjected to passive movements for periods ranging from 1 to 6
h, at a rate of 300 movements per minute. Rais (16) exercised rabbits for 0.5 to 6 h with a frequency of 150 movements per minute and used electrical stimulation of the sciatic nerve to accomplish an active contraction. The latter author demonstrated slight edema in muscle, paratenon, and mesotenon after 0.5 h of exercise. The degree of these changes correlated to the duration of the exercise. Perivascular accumulations of polymorphonuclear leukocytes and an increased number of fibroblasts in the paratenon and mesotenon were noted on histopathological examination of specimens from rabbits killed 4 days after an exercise period of 6 h. In this study, an experimental model in the rabbit was designed and tested in an attempt to standardize the conditions related to the development of chronic Achilles paratenonitis with tendinosis. MATERIAL AND METHODS
The experiment was carried out on 13 New Zealand white rabbits of both sexes aged from 6 to 9 months. The animals were caged all of their lives.
Address correspondence and reprint requests to Dr. C. Backman, Department of Hand Surgery, University Hospital of Umel, S-901 85 Umel, Sweden.
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Their weights ranged from 2.5 to 4 kg. Before and after exercise, the animals were kept in ordinary cages allowing good freedom of movement and had free access to food and water. Throughout the experiment, the rabbits were kept under anesthesia, induced by intramuscular injection of fentanylfluanison (0.3 ml/kg) and diazepam (2 mg/kg). This was followed by another injection of fentanylfluanison (0.15 ml/kg) 30 min later. A kicking machine (Fig. 1) was used for passive flexions and extensions of the ankle joint. The movements were produced by a pneumatic piston, where the range of movement could be altered. The movement was set to 9.5 cm, giving a range of motion in the ankle of 55-65", of which 20-25" was dorsiflexion and 3540" was plantar flexion. In order to attain this angular movement, a band was tied around the thigh of the rabbit to restrict the motion of the hip and knee. The piston was attached to the right foot. The left leg served as control and was at rest throughout the experiment. The machine was constructed for the simultaneous exercising of three rabbits. An active contraction was induced during the plantar flexion phase by electrical stimulation via surface electrodes (Pediatric electrodes 40 426A, Hewlett Packard, Andover, MA, U.S.A.) placed 2 cm apart over the triceps surae muscles. This stimulation was synchronized with the plantar flexion movement of the piston by a microswitch, which
5 A
7
1
trigged the stimulator unit (Disa stimulator Type 14E 10, Disa Elekronik A / S , Herlev, Denmark). The stimulator was set to generate an impulse train of 92 ms (impulse duration of 0.2 ms at 18 ms intervals), giving rise to six pulses per train. The pattern of the stimulus was set after pilot experiments. The muscle contraction was triggered 130 ms before the piston was reversed. The time between muscle stimulation and peak tension (6) makes the triceps surae muscles contract during the prestrech and the initiation of the passive dorsiflexion movement. The electrical stimulus was controlled by oscillometer. We used the same setup in all experiments, except for the amplitude, which ranged between 35 and 50 V, depending on the individual contraction response. Throughout the experiment, the contraction response was controlled and if necessary the amplitude was adjusted during the exercise in order to achieve a constant contraction. In a pilot study, five rabbits were exercised for 1 h three times a week for 1 to 6 weeks and the tendon was palpated regularly. The frequency of the flexion and extension movements was set to 150 per minute. Following 6 weeks of exercise, inflammatory alterations in the paratenon was the major histopathological finding. Lymphocytes predominated among the inflammatory cells. The signs of degeneration in the tendon, with variation in staining affinity and varying thickness of the fibrillar components, were present but only to a minor extent. Based on clinical and histopathological findings, 2 h of exercise three times a week for 5 to 6 weeks was used for this study. After the animals were sacrificed (the day after the last exercise), the Achilles tendons from both legs were excised and placed in formalin. The specimen was kept in formalin for 2-3 days. After dehydration in alcohol, the tendon was embedded in paraffin and longitudinally cut (section thickness of 5-6 Fm). The sections were stained with hematoxylin-eosin.
RESULTS
FIG. 1. (A) Rabbits positioned in kicking apparatus. (1) DlSA stimulator unit. (2) Switch for individual adjustment of amplitude. (3) Pneumatic piston. (4) Band restricting the movement of hip and knee. (B) Closeup view of hindfoot and pneumatic piston. Microswitches (5) regulate the motion length of the piston and switch (6) triggers the DlSA stimulator unit. Surface electrodes placed on the triceps surae muscles (7).
J Orthop Res, Vol. 8, No. 4, 1990
Throughout the experimental period, the animals were in generally good condition. After 4 weeks of exercise, palpation of the exercised leg indicated that all animals had irregular thickening over the Achilles tendon. At this time, most animals demonstrated palpable nodules 0.5 to 1 cm above the insertion into the calcaneus. Light microscopic examination disclosed a normal histological appearance of tendon and paratenon in the control legs (Figs. 2A and 3A).
EXPERIMENTAL CHRONIC ACHILLES PARA TENONITIS The tendons of the exercised legs exhibited degenerative changes of varying severity and distribution. Most often, these changes were localized to the central portion of the tendons (Fig. 2B). The staining affinity varied in different portions of the tendons, and the fibrillar components showed varying thickness (Fig. 2C, 2D). The nuclei of the ten-
543
dons exhibited slightly varying size, contour, and staining affinity. Fibrillation was occasionally encountered in the tendons (Fig. 3B). The paratenon of the exercised legs was thickened (Figs. 4A-D and 5A) with the occurrence of an increased number of fibroblasts (Figs. 3B and 4AD). The number of capillaries was also increased
A.B
FIG. 2. (A) Low-power micrograph demonstrating a normal appearance of tendon and thin paratenon (to the right). Control leg, 6 weeks. Hematoxylin-eosin, x25. (6)Low-power micrograph showing degenerative changes with varying staining affinity in the tendon, and thickened paratenon (to the right). Exercised leg, 6 weeks. Hematoxylin-eosin, ~ 3 7 (C) . Micrograph showing degenerative tendon. Exercised leg, 5 weeks. Hematoxylin-eosin, ~ 1 5 0(D) . Higher magnification demonstrating degenerative alterations in tendon with varying staining affinity. Exercised leg, 6 weeks. Hematoxylin-eosin, ~237.
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FIG. 3. (A) Normal tendon and paratenon. Control leg, 6 weeks. Hernatoxylin-eosin, x375. (B) Micrograph demonstrating fibrillation in tendon, dilated capillary (C) containing erythrocytes, and increased number of firoblasts (F)in paratenon. Exercised leg, 5 weeks. Hematoxylin-eosin, x375.
(Figs. 4C and 5A), and these vessels were frequently dilated (Fig. 5A) and filled with erythrocytes (Fig 3B). Additional changes in the paratenon of the exercised legs were edema (Figs. 5A, 5B) and infiltration of inflammatory cells (Figs. 5B-D). Lymphocytes predominated in the inflammatory cell idiltrates, but plasma cells and granulocytes were also encountered. A varying number of eosinophils could be observed, in addition to neutrophils. Histiocytes were also found. A distinct perivascular distribution of inflammatory cells was a common finding (Figs. 5D). Table 1 shows a semiquantitative assessment of the histopathological changes observed in each experimental animal. DISCUSSION
This study demonstrates an animal model for inducing histopathological changes associated with chronic Achilles paratenonitis and paratenonitis with tendinosis, i.e., degenerative alterations of tendon, inflammatory cells, increased number of capillaries, as well as edema and fibrosis in the paratenon (15). Although the extent of histopathological change varied among the experimental animals, all of these animals exhibited alterations both
J Orthop Res, Vol. 8, No. 4, 1990
in tendon and paratenon as demonstrated in a semiquantitative manner in Table 1. Previous experimental studies have focused upon the acute Achilles paratenonitis (14,16). In order, however, to correlate to the prolonged overuse situation in humans, predisposing for tendon ruptures (4,5,11,17), a more extended period of stimulus imposed on the tendon is necessary (2). The choice of this protocol was based on preliminary experiments indicating that 1 h of exposure (three times per week for 6 weeks) to exercise brought about inflammatory changes in the paratenon but merely minor signs of degeneration within the tendon (Backman et al., unpublished data). Also, pilot experiments showed that 4 weeks of exercise gave rise to palpatory changes, i.e., nodules around the tendon and diffuse thickening of the soft tissues that are in concert with the observations reported by Kvist et al. (8). In order to mimic the etiological factors in humans, the rabbit tendons were subjected to simultaneous electrical muscle stimulation and traction of the Achilles tendon. This regimen has previously been shown to be the most successful way of inducing acute inflammatory response in the Achilles paratenon (16). The stimulation protocol, giving impulse trains of 92 ms duration, indicates the necessity of a more extended contraction, as opposed to
EXPERIMENTAL CHRONIC ACHILLES PARA TENONITIS
545
C.D
FIG. 4. (A) Low-power micrograph showing markedly thickened paratenon (P), increased number of capillaries (C) and adipose tissue (A). The tendon is seen to the left. Exercised leg, 6 weeks. Hematoxylin-eosin, ~ 4 3(6) . Higher magnification of thickened paratenon showing numerous fibroblasts (F). Adipose tissue (A) and tendon (T) are also seen. Exercised leg, 6 weeks. Hematoxylin-eosin, x95. (C) Micrograph showing thickened paratenon with increased number of capillaries and fibroblasts. Exercised leg, 6 weeks. Hematoxylin-eosin, x95. (D) Micrograph showing paratenon with increased number of fibroblasts and fibrils. Exercised leg, 5 weeks. Hematoxylin-eosin, x375.
a single twitch. The stimulation during the plantar flexion phase creates a contraction of the myotendinous tissue while in a shortened position and through the initiation of the dorsiflexion movement (6). This is in line with recent studies demonstrating
that during running as well as normal stride, a prestretch of the gastrocnemii occurs prior to foot placement when landing (13). To induce hyperfunction, we applied the same kicking frequency as that implemented to cause
J Orthop Res, Vol. 8, No. 4, 1990
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FIG. 5. (A) Micrograph demonstrating wavy appearance in tendon (T) and markedly thickened paratenon with dilated capillaries (C),edema (E), and fibrosis. Exercised leg, 5 weeks. Hematoxylin-eosin, x60. (B) Higher magnification showing edema (E) and inflammatory cells (I), including lymphocytes and granulocytes, and some fibroblasts. Exercised leg, 5 weeks. Hematoxylineosin, x375. (C) A rich number of inflammatory cells, mainly lymphocytes, but also some plasma cells and granulocytes, is interspersed among connective tissue fibrils and small edematous areas in paratenon. Exercised leg, 5 weeks. Hematoxylineosin, x375. (D)Micrograph demonstrating perivascular infiltration of inflammatory cells in adipose tissue close to a tendon. Exercised leg, 5 weeks. Hematoxylin-eosin, ~ 2 3 7 .
acute paratenonitis (16). Due to the fact that this is a survival experiment, we modified the stimulation technique and employed transcutaneous electrical stimulation of the muscles, as opposed to the indirect sciatic nerve stimulation used by Rais (16). A higher stimulation voltage is necessary to proJ Orthop Res, Vol. 8, No. 4, 1990
duce an active contraction when applying transcutaneous muscle stimulation as compared to nerve stimulation. The transcutaneous stimulation is, however, easily reproduced, noninvasive, and convenient for repeated application. It may be argued that the transcutaneous stimulation per se could
547
EXPERIMENTAL CHRONIC ACHILLES PARATENONITIS TABLE 1. Semiquantitative assessment of histopathological changes in each experimental animal Tendon Animal no. 6 7 8 9 10 11 12 13 14 15
16 17 18
0
=
Paratenon
Staining affinity
Nuclear appearance
Fibrillar appearance
2 1 3 3 1 0 3 2 0 3 2 3 3
1 1 2 1 2 2 2 3 0 3 2 1 2
1 2 3 2 2 1 3 2 1 2 3
1 2 1 3 3 0 2 1 3 2
1
2 3
no change, 1
=
slight change, 2
3 =
Thickness
3
moderate change, and 3
cause an inflammatory reaction in the tendon. There are two major objections to such an assumption. First, the electrodes were located at some distance from the tendon. Second, changes related to electrical stimulation appear to be confined to muscle tissue (1,12). Our findings are identical to those reported in biopsy material from professional runners and joggers with sustained Achilles tendon complaints, admitted for surgery after months or years of nonbeneficia1 conservative treatment (2). The described biopsy material shows paratendineal thickening, edema, proliferation of fibroblasts, formation of new connective tissue, infiltrates of inflammatory cells, and proliferation of small blood vessels in the paratenon (7). The inflammatory changes in paratenon are often combined with areas of focal degeneration within the tendon, indicating paratenonitis with tendinosis (2). The presence and degree of edema and polymorphonuclear leukocytes is a sign of acute peritendinis. This result is explained by the fact that the animals were sacrificed the day after their last exercise bout. In conclusion, this model provides a tool for the further understanding of the mechanisms involved in the development and possibly in the treatment of chronic Achilles paratenonitis with tendinosis. Acknowledgment: This work has been supported by grants from the Research Council of the Swedish Sports Federation and the Medical Faculty, Umei University.
REFERENCES 1. Cabric M, Appell HJ, Resic A: Effects of electrical stimula-
tion of different frequencies on the myonuclei and fiber size in human muscle. Int J Sports Med 8:323-326, 1987
=
Occurrence of fibrosis
Occurrence of edema
1 2 I 3 2 0 2 2 2 2 3 1 2
0 1 0 1 2 3
0 1 0 0
1 2 3
Capillaries
Inflammation
2 2 3 1 3 3 1 2 2 1 2 3 2
3 2 3 2 3 3 2 1 0 1 3 2 2
marked change. 2. Clancy WG, Neidhart D, Brand RL: Achilles tendonitis in runners: A report of five cases. Am J Sports Med 4:46-57, 1976 3. Clement DB, Taunton JE, Smart GW: Achilles tendinitis and peritendinitis: Etiology and treatment. Am J Sports Med 12:179-184, 1984 4. Davidsson L, Salo M: Pathogenesis of subcutaneous tendon ruptures. Acta Chir Scand 135:20%212, 1969 5. Fox JM, Blazina ME, Jobe FW, Kerlan RK, Carter VS, Shields CL, Carlson GJ: Degeneration and rupture of the Achilles tendon. Clin Orthop 107:221-224, 1975 6. Freund HJ: Motor unit and muscle activity in voluntary motor control. Physiol Rev 63:387436, 1983 7. Kvist M, Jozsa L, Jarvinen M, Kvist H: Chronic achilles paratenonitis in athletes: A histological and histochemical study. Pathology 19:1-1 1 , 1987 8. Kvist H, Kvist M: The operative treatment of chronic calcaneal paratenonitis. J Bone Joint Surg [Br] 62:353-357, 1980 9. Leach RE, DiIorio E, Harney RA: Pathologic hindfoot conditions in the athlete. Clin Orthop 177:116-121, 1983 10. Leach RE, James S, Wasilewski S: Achilles tendinitis. Am J Sports Med 9:93-98, 1981 1 1 . Ljungkvist R: Subcutaneous partial rupture of the Achilles tendon. Acta Orthop Scand Suppl 113:1-86, 1968 12. Myrhage R, Hudlicka 0: Capillary growth in chronically stimulated adult skeletal muscle as studied by intravital microscopy and histological methods in rabbits and rats. Microvasc Res 16:73-90, 1978 13. Nilsson J, Thorstensson A, Halbertsma J: Changes in leg movements and muscle activity with speed of locomotion and mode of progression in humans. Acta Physiol Scand 123:457-476, 1985 14. Obolenskaja AJ, Goljanitzki JA: Die serose Tendovaginitis in der Klinik und im Experiment. Dtsch Z Chir 201:388-399, 1927 15. Puddu G, Ippolito E, Postacchini F: A classifcation of achilles tendon disease. Am J Sports Med 4:145-150, 1976 16. Rais 0:Heparin treatment of peritenomyosis (peritendinitis) crepitans acuta. Acta Chir Scand Suppl268: 1-88, 1961 17. Skeoch DU: Spontaneous partial subcutaneous ruptures of the tendo achilles. Am J Sports Med 9:20-22, 1981 18. Snook GA: Achilles tendon tenosynovitis in long distance runners. Med Sci Sporrs 4:155-158, 1972
J Orthop Res, Vol. 8, No. 4, 1990
