Hyaluronic acid modulates cell proliferation unequally in intrasynovial and extrasynovial rabbit tendons in vitro

As tendons differ in biochemical composition and cellular capacities, we have compared dose response effects of hyaluronic acid on cell proliferation and synthesis of matrix components in intermediate and proximal segments of intrasynovial deep flexor tendons and extrasynovial peroneus rabbit tendons in vitro. Compared with matched control tendons, hyaluronic acid inhibited cell proliferation in intermediate and proximal intrasynovial flexor tendon segments at the concentrations of 0.1-2.0 mg/ml and 0.5-2.0 mg/ml respectively, but in extrasynovial tendon segments only at the concentration of 0.5 mg/ml. Hyaluronic acid did not affect synthesis of proteoglycan, collagen and non-collagen protein in either type of tendon. These results show that hyaluronic acid modulates cell proliferation unequally in intra- and extrasynovial tendons without affecting the synthesis of matrix components in the two types of tendons, indicating differential hyaluronic acid sensitivity and a possible mechanism of action.     

Dose-related cellular effects of platelet-derived growth factor-BB differ in various types of rabbit tendons in vitro

Cellular responsiveness to growth factors that can affect tendon healing may be site-specific. We have compared the dose-response effects of platelet-derived growth factor-BB (PDGF-BB) on proteoglycan, collagen, noncollagen protein and DNA synthesis between intrasynovial intermediate and proximal segments of deep flexor tendons, and extrasynovial peroneal tendons of rabbits during short-term cultures. PDGF-BB stimulated matrix and DNA synthesis of the three types of tendon segments in a dose-dependent manner in the range from 0.1 to 100 ng/mL. PDGF-BB stimulated collagen synthesis and noncollagen protein synthesis (calculated from LogED 50 ) in proximal intrasynovial tendon segments more than in extrasynovial peroneal tendon segments, and DNA synthesis less in proximal than in intermediate intrasynovial tendons. However, the estimated maximal stimulations (E max ) by PDGF-BB were similar in the three types of tendon segments. These findings show that PDGF-BB stimulates DNA and matrix synthesis differently in various types and regions of tendons during short term explant culture and suggests that there may be differences in cellular responsiveness during tendon healing.     

Dose-related cellular effects of platelet-derived growth factor-BB differ in various types of rabbit tendons in vitro

Cellular responsiveness to growth factors that can affect tendon healing may be site-specific. We have compared the dose-response effects of platelet-derived growth factor-BB (PDGF-BB) on proteoglycan, collagen, noncollagen protein and DNA synthesis between intrasynovial intermediate and proximal segments of deep flexor tendons, and extrasynovial peroneal tendons of rabbits during short-term cultures. PDGF-BB stimulated matrix and DNA synthesis of the three types of tendon segments in a dose-dependent manner in the range from 0.1 to 100 ng/mL. PDGF-BB stimulated collagen synthesis and noncollagen protein synthesis (calculated from LogED50) in proximal intrasynovial tendon segments more than in extrasynovial peroneal tendon segments, and DNA synthesis less in proximal than in intermediate intrasynovial tendons. However, the estimated maximal stimulations (Emax) by PDGF-BB were similar in the three types of tendon segments. These findings show that PDGF-BB stimulates DNA and matrix synthesis differently in various types and regions of tendons during short term explant culture and suggests that there may be differences in cellular responsiveness during tendon healing.     

Dose-related cellular effects of platelet-derived growth factor-BB differ in various types of rabbit tendons in vitro

Cellular responsiveness to growth factors that can affect tendon healing may be site-specific. We have compared the dose-response effects of platelet-derived growth factor-BB (PDGF-BB) on proteoglycan, collagen, noncollagen protein and DNA synthesis between intrasynovial intermediate and proximal segments of deep flexor tendons, and extrasynovial peroneal tendons of rabbits during short-term cultures. PDGF-BB stimulated matrix and DNA synthesis of the three types of tendon segments in a dose-dependent manner in the range from 0.1 to 100 ng/mL. PDGF-BB stimulated collagen synthesis and noncollagen protein synthesis (calculated from LogED 50 ) in proximal intrasynovial tendon segments more than in extrasynovial peroneal tendon segments, and DNA synthesis less in proximal than in intermediate intrasynovial tendons. However, the estimated maximal stimulations (E max ) by PDGF-BB were similar in the three types of tendon segments. These findings show that PDGF-BB stimulates DNA and matrix synthesis differently in various types and regions of tendons during short term explant culture and suggests that there may be differences in cellular responsiveness during tendon healing.     

Differential effects of insulin-like growth factor-I on matrix and DNA synthesis in various regions and types of rabbit tendons

Tendon healing and integration of tendon grafts may be site or donor specific. To determine if differences exist in sensitivity to growth factors that have the potential to influence tendon repair, we compared the effects of recombinant human insulin-like growth factor-I on various types of tendon segments. The dose response effects on proteoglycan, collagen, noncollagen protein, and DNA synthesis were investigated in short-term explant cultures of intrasynovial intermediate and proximal segments of deep flexor tendons, extrasynovial segments of deep flexor tendons, and Achilles tendons of rabbits. The four different types of tendon segments cultured in media without recombinant human insulin-like growth factor-I synthesized similar amounts of each of the matrix components. Intrasynovial proximal segments synthesized 15 times less DNA than other tendon segments. Recombinant human insulin-like growth factor-I stimulated matrix and DNA synthesis of all tendon segments in a dose-dependent manner in intervals from 10 to 1,000 ng/ml. The potency (LogED₅₀) of the stimulation did not differ between the segments. The estimated maximal stimulation (E_max) of proteoglycan synthesis by recombinant human insulin-like growth factor-I was higher, and of collagen and noncollagen protein synthesis was lower, in intrasynovial proximal segments as compared with that of the other types of segments. In contrast, the estimated maximal stimulation of DNA synthesis by recombinant human insulin-like growth factor-I was 6-fold higher than controls in all types of tendons. These findings demonstrate differences in mitotic capacity between anatomical regions of tendons during culture without recombinant human insulin-like growth factor-I and in matrix synthesis after stimulation with it.     

Collagen synthesis during primate flexor tendon repair in vitro

The concept that flexor tendons have the intrinsic capacity to participate actively in the repair process following laceration has been developed in recent years as the result of experimental studies from numerous laboratories. However, the role of the outer/epitenon and inner/endotenon cell populations with regard to protein synthesis is still controversial. The purpose of this study was to investigate the respective participation of these tendon fibroblast populations in the synthesis of the collagen matrix during in vitro repair of the flexor tendon from nonhuman primates, utilizing immunohistochemical techniques and a Type I procollagen antibody. Zone II profundus flexor tendon segments were obtained from young adult Macaca nemestrina monkeys. One centimeter segments were cultured, either with or without a transverse laceration across 90% of the midsection of the tendon segment. Frozen sections of the cultured tendon segments were reacted with the (mouse monoclonal) Type I procollagen antisera. At all times of culture of the nonlacerated tendon segment, only a few of the epitenon cells along the surface of the tendon and distant from the cut end stained positively for active collagen synthesis. In the lacerated segments, and as early as 9 days of culture and repair, the procollagen reaction product was starting to appear in those cells of the outer epitenon cell layer. These studies support the concept that the inner fibroblasts do actively participate in collagen production. However, it now also appears that a significant degree of collagen synthesis during tendon repair resides in the outer/epitenon layer of cells enveloping the tendon segment, and that the repair response of the flexor tendon in vitro is proportional to the degree of injury.     

Tenascin-C in tendon regions subjected to compression.

Tendon regions subjected almost exclusively to tension differ from regions subjected to high levels of compression as well as tension. Regions not exposed to compression consist primarily of spindle-shaped fibroblasts surrounded by densely packed longitudinally oriented collagen fibrils formed principally from type-I collagen. In contrast, regions subjected to compression have a fibrocartilagenous structure and composition: they consist of spherical cells surrounded by a matrix containing hyaline cartilage proteoglycans (aggrecan) and type-II collagen as well as type-I collagen. Reducing their adhesion to the matrix may help cells in the latter regions establish and maintain a spherical shape and minimize their deformation when the tissue is subjected to mechanical stress. We hypothesized that expression of tenascin-C, an anti-adhesive protein, is part of the adaptation of tendon cells to compression that helps establish and maintain fibrocartilagenous regions. To test this hypothesis, we compared segments of bovine flexor tendons subjected to repetitive compression (distal) with segments that are not subjected to compression (proximal) to determine whether they differed in tenascin-C content and expression. RNA and protein analyses showed that tenascin-C expression was elevated in the distal tendon. Tendon cells from the distal segment expressed more tenascin-C mRNA than did cells from the proximal segments for as long as 4 days in cell culture, indicating that increased tenascin-C expression is a relatively stable feature of the distal cells. Moreover, purified tenascin-C inhibited the attachment of cultured tendon cells to fibronectin. These observations support the hypothesis that tenascin-C expression is a cellular adaptation to compression that helps establish and maintain fibrocartilagenous regions of tendons by decreasing cell-matrix adhesion.     

Division of flexor tendons causes progressive degradation of tendon matrix in rabbits

Deep flexor tendons of 30 rabbits were divided at the ankle level. the effects of unloading on the synthesis and content of matrix components, the synthesis of DNA, and dry weight were investigated. the reaction of the fibrocartilaginous and non-fibro-cartilaginous segments were separately analyzed.

The ability of the tendons to synthesize collagen during short-term culture and the contents of matrix components decreased inversely to the time of unloading. 12 weeks following division, the fibro cartilaginous segments had lost 2/5 of their dry weight, 2/3 of proteoglycan and 1/3 of collagen and non-collagen protein content. Less pronounced losses were observed in the non-fibrocartilaginous segments. A transient increase in cell proliferation in both types of segments was found. These findings indicate that divided flexor tendons undergo a progressive degradation, which may have implications for delayed suture of deep flexor tendon injuries.     

Regional differences in matrix formation in the healing flexor tendon

In 20 white leghorn chickens, the effect of proximity (proximal vs. distal) to the repair site and the effect of sheath excision versus sheath repair on biochemical matrix formation in the healing zone 2 flexor tendon were studied. In 14 animals, the profundus tendon of the long toe on both feet was lacerated and repaired in zone 2. In the right foot, the sheath was repaired, and on the left foot, the sheath was excised. Segments proximal and distal to the laceration site were studied at six weeks postrepair compared to control segments. Sheath excision versus sheath repair had no effect on the net matrix formation of the healing flexor tendon. A higher DNA content and lower hydroxyproline and hexosamine contents were present in the healing tendons, indicating the healing process was not complete at six weeks. Sheath excision versus sheath repair had no effect on net matrix formation in the healing tendon. There is no difference in DNA content or glycosaminoglycan content in the proximal versus distal segments. There was a significantly greater net hydroxyproline content in the proximal segment versus the distal segment of the healing tendon, which was not present in similar control segments. These results support the concept that the zone 2 flexor tendon does not respond to injury as a homogeneous structure.     

Dehydration inhibits matrix synthesis and cell proliferation. An in vitro study of rabbit flexor tendons

Segments of the deep flexor tendon of the rabbit were exposed to air; the effects of dehydration on in vitro synthesis of proteoglycan, collagen, noncollagenous protein, and cell proliferation were compared with tendon segments that were kept moist with physiologic saline. After 20 min of exposure to air, the tendons lost half and after 40 min all of their ability to synthesize matrix components and to proliferate, whereas irrigated tendons remained viable during the entire experiment.     

Long-term explant culture of rabbit flexor tendon: effects of recombinant human insulin-like growth factor-I and serum on matrix metabolism

The effects of human recombinant insulin-like growth factor-I (rhIGF-I, 50 ng/ml) on matrix metabolism in the deep flexor tendon from the tendon sheath region of the rabbit were studied in explants cultured for 3 weeks. Tendon segments cultured in medium supplemented with fetal calf serum (FCS) exhibited proliferation of the superficial cell layers. Synthesis of proteoglycan and non-collagen protein (NCP) increased threefold during the first week and remained elevated during the next 2 weeks of culture in medium supplemented with rhIGF-I or FCS, but not in medium without supplements (bovine serum albumin, BSA). The estimated halflife (t1/2) for elimination of newly labeled proteoglycans from the tendon explants ranged from 5.1 to 8.5 days and from 4.9 to 6.8 days for NCP in supplemented medium. Presence of rhIGF-I or FCS did not affect degradation of matrix as compared with BSA. The total hexosamine content per tendon segment was stable during the culture period, but the non-collagen protein content decreased by 25%. Collagen synthesis decreased to 10% of the initial level after 3 weeks in supplemented medium, but to 3% in unsupplemented medium. There was no measurable turnover of collagen in explants cultured in either medium, and the collagen content remained unchanged. Our results suggest that rhIGF-I, as well as FCS, stimulates matrix synthesis but does not influence matrix turnover in rabbit flexor tendon explants in long-term culture as compared with medium without supplements. We conclude that rhIGF-I may be used as a defined growth-promoting factor in serum-free media and may be of importance in tendon healing.     

Distal anatomical relationship of the flexor hallucis longus and flexor digitorum longus tendons

BACKGROUND: The distal attachment of the flexor hallucis longus (FHL) tendon with the flexor digitorum longus (FDL) tendon varies antomically. The presence of a strong link between the two tendons can preserve distal function if one of the tendons is used for transfer. METHODS: Twenty-four cadaver legs were dissected, and the distal relationship of the FHL tendon with the FDL tendon was analyzed. The width of the tendons and their attachments were measured to the closest 0.5 mm. RESULTS: Three different configurations were found. In type 1, a tendinous slip branched from the FHL to the FDL (10 of 24 feet). In type 2, a slip branched from the FHL to the FDL and another from the FDL to FHL (10 of 24). In type 3, no attachment was present (four of 24). In four cadavers the attachment was different in the right and left feet. CONCLUSION AND CLINICAL RELEVANCE: The absence of a cross connection between the two tendons in the foot may be more frequent than previously reported. Three configurations of the anatomical relationship of the distal FDL to FHL tendons were found in this study with a small sample size. Based on these findings, to preserve the distal function of the FDL after transfer of the proximal FDL tendon, routine tenodesis should be done or a wider exposure and tenodesis in type 3 variations.     

Variations of extensor pollicis brevis tendon in Indian population: A cadaveric study and review of literature

ObjectivesVariations of the tendons of the first dorsal compartment of the wrist may be one of reasons of treatment failure and recurrence in De Quervain's tenosynovitis (DQT). The present cadaveric study was designed to look into the variations of the Extensor pollicis brevis (EPB) tendon in Indian population.MethodsSeventy-seven formaldehyde-fixed cadaveric upper limbs of Indian origins were dissected to observe the number of EPB tendons and its variations.ResultsThe EPB muscle was found to be absent in one hand (1.3%). The EPB muscle was found with single tendon, two tendons and three tendons in 73 limbs (94.8%), 2 limbs (2.6%) and one limb (1.3%) respectively. The muscle originated from the posterior surface of the radius and the adjacent interosseous membrane. The EPB muscle with single tendon was found to be inserted into the distal part of dorsal surface of the proximal phalanx of the thumb in 44 limbs (57.1%). In limbs with bitendinous EPB, the tendon slips were inserted into the base of proximal phalanx and into the base of distal phalanx of the thumb. An Osseo-fibrous septum separating EPB from Abductor Pollicis Longus (APL) was observed in 45 limbs (58%).ConclusionEPB in first extensor compartment of Indians is usually monotendinous. It mostly inserts into the distal part of dorsal surface of proximal phalanx of thumb and into the base of distal phalanx. In majority of the wrists, one may find an osseofibrous ridge separating EPB from APL. These anatomical variations may be helpful to guide proper treatment in de Quervain’s tenosynovitis.     

Recombinant human insulin-like growth factor-I stimulates in vitro matrix synthesis and cell proliferation in rabbit flexor tendon

Flexor tendons have an intrinsic ability for repair, with a capacity to metabolize matrix components and to proliferate. To identify factors with the potential of affecting those abilities, the effects of recombinant human insulin-like growth factor (rhIGF-I), insulin and fetal calf serum (FCS) on the synthesis of proteoglycan, collagen, and non-collagen protein and cell proliferation were investigated in short-term explant cultures of the deep flexor tendon of the rabbit. Matrix synthesis and cell proliferation were stimulated dose dependently by rhIGF-I at doses between 10 and 250 and at 10-100 ng/ml, respectively, by insulin at 250-5,000 ng/ml, and by FCS at 2-15%. Estimated maximal stimulation (Emax) of up to three times the control value was observed with rhIGF-I at 250 ng/ml. Maximal stimulation was observed at 5,000 ng/ml with insulin, and FCS at 15%. rhIGF-I was more potent than insulin in stimulating protein synthesis and cell proliferation. The Emax of stimulation of proteoglycan and collagen synthesis by rhIGF-I were two times that of FCS, and the Emax of cell proliferation by FCS was twice that of rhIGF-I. Growth factors thus have the ability to stimulate matrix synthesis and cell proliferation in rabbit flexor tendon. This provides a rationale for further studies on the role of growth factors in flexor tendon healing in humans.     

The fate of tendon grafts used to reconstruct the digital annular pulleys

An experimental morphological study on the reconstruction of the digital annular pulleys has been carried out in dogs. The segment corresponding to zones 1 and 2 of the flexor apparatus of the 2nd and 5th digits of the left forepaw was chosen for the experiment. The whole flexor apparatus was resected and a single digital pulley (A 2) was reconstructed, using segments of the animals own deep flexor tendon. A length of silicone rubber tube was used as tendon spacer. The new pulleys showed marked degeneration of the collagen fibres and thinning which increased with time and may be the cause of decrease in strength. A layer of mesothelial cells with secretory properties developed at the interface between the tendon graft and the spacer.     

Popliteoperoneal in situ bypass using the small saphenous vein enables revascularization with low trauma

In patients with diabetes, a popliteocrural vein bypass frequently must be linked to the distal peroneal artery. To reduce trauma to the ischemically damaged tissue, we used a dorsal approach to the peroneal artery. With the patient prone, a incision parallel to the posterolateral margin of the Achilles tendon is made. After the deep crural fascia and flexor hallucis longus are split, access to the peroneal artery is easily obtained. The second segment of the popliteal artery is exposed in the popliteal cavity. The small saphenous vein is left in situ and anastomosed with the arteries after proximal and distal preparation and valvulotomy.     

The effects of reduced oxygen tension on cell proliferation and matrix synthesis in synovium and tendon explants from the rabbit carpal tunnel: an experimental study in vitro.

Local ischemia may play an important role in the development of tendon degeneration as well as entrapment neuropathies. In order to investigate the cellular effects of hypoxia on tendon and synovial tissue from the carpal canal, dose response effects of oxygen on cell proliferation and synthesis of matrix components were examined in segments of synovial and flexor digitorum profundus tendon from the carpal tunnel of rabbits during short term culture. Explants were incubated in airtight containers flushed with either 0%, 1%, 3%, 20% O2 plus 2% CO2 and N2 to balance and labeled with either 3H-thymidine or 3H-proline and 35S-sulfate. Cell proliferation was significantly inhibited by hypoxia in synovium but not in tendon (P = 0.03). In parallel, the synthesis of non-collagenous proteins was significantly reduced in synovium but not in tendon (P = 0.006). In both tissues hypoxia significantly inhibited collagen synthesis. On the other hand, hypoxia had no significant effect on the synthesis of new proteoglycans as determined by 35S-sulfate incorporation. Hypoxia can inhibit cell proliferation and alter synthesis of matrix components in synovial tissue, but may only have minor effects on non-collagen protein synthesis in tendon explants from the carpal canal of rabbit forepaws.     

Der popliteoperoneale In-situ-Bypass mit der V. saphena parva

In diabetics, a popliteocrural vein bypass frequently has to be linked to the distal fibular artery. To reduce trauma to the already ischaemically damaged tissue, dorsal access to the fibular artery was selected. With the patient in the prone position, the fibular artery is exposed by a premalleolar incision lateral to the Achilles tendon. Splitting of the deep crural fascia and the flexor hallucis longus allows access to the fibular artery without problem. The second segment of the popliteal artery is exposed in the popliteal cavity. The small saphenous vein is left in situ and anastomosed to the arteries after proximal and distal preparation and valvulotomy. With dorsal access, the distal fibular artery can be exposed quickly and with little trauma. Tissue trauma is further minimized by the in-situ bypass routing of the small saphenous vein. Prerequisites are a good flow to the second segment of the popliteal artery and a usable small saphenous vein.