Laurecir Gomes

Obesity Affects Collagen Fibril Diameter and Mechanical Properties of Tendons in Zucker Rats

Obesity is currently considered to be a world epidemic and one of the major public health problems in many countries, whose incidence is increasing at alarming rates. Genetically obese Zucker rats are used as a model of obesity and were employed in the present study. Tendons transmit contractile force from muscles to bone, thus permitting articular movement. The objective of our study was to analyze the ultrastructural, biochemical, and biomechanical alterations that occur in the deep digital flexor tendon of obese Zucker rats compared to lean animals. Ultrastructural analysis showed differences in collagen fibril diameter distribution and mass-average diameter between obese and lean animals. Regarding mechanical parameters, there was a significant difference in maximum displacement and strain. Hydroxyproline content was higher in obese animals. In view of the excess weight and peculiar conditions to which the tendon of obese animals is submitted, we concluded that obesity provokes alterations in the composition and organization of tendon extracellular matrix components. These alterations might be related to organizational and structural modifications in the collagen bundles, influencing the mechanical properties of the tendon and the progression to a pathological state.

Structural and biochemical alterations during the healing process of tendons treated with Aloe vera

AIMS The tendon is composed of highly organized collagen fibers that form a complex supramolecular structure. After lesions, the organization and composition of the tendon are not completely restored. Our purpose was to evaluate if the application of Aloe vera improves tendon healing, considering the effectiveness in the stimulus of collagen synthesis. MAIN METHODS The calcaneal tendon of male Wistar rats was partially transected with subsequent topical application of A. vera ointment at the injury. The animals were separated into groups with tendons treated with the A. vera extract for 7days and excised on the 7th, 14th and 21st days after surgery; control rats received only ointment base without plant extract. KEY FINDINGS Morphological analysis using polarization microscopy showed that the entire tendon undergoes a remodeling process, with disorganized collagen fibers by days 7 and 14 in plant-treated and non-treated groups and with a higher birefringence in tendons of the plant-treated group on the 21st day. A higher concentration of hydroxyproline was found in plant-treated tendons on days 7 and 14 compared with their controls. Western blots showed lower amounts of type I collagen in the plant-treated group on day 14 compared with the control. MMP-9 diminished 14days after lesion and the active isoform of MMP-2 increased on day 21 in plant-treated groups. SIGNIFICANCE The present study indicates a beneficial effect of A. vera in the tissue reorganization in the transected region of the tendon 21days after injury and is supported by an increase of active MMP-2.

Biochemical and Biomechanical Analysis of Tendons of Caged and Penned Chickens

Chickens were divided into two groups, one caged and the other penned. Superficial digital flexor tendons from penned chickens showed greater tensile strength, withstanding a greater strain before rupture than tendons from caged chickens. The tensile region of tendons from penned chickens showed more swelling in acetic acid and a higher hydroxyproline concentration compared with caged chickens, indicating the presence of large collagen amounts in the former. The tensile region of penned chickens presented higher glycosaminoglycan concentrations than the same region of caged chickens. For both groups, the predominant glycosaminoglycan in the compression regions was chondroitin sulfate, whereas dermatan sulfate was found in the tensile regions. N-terminal analysis identified the small proteoglycans fibromodulin and decorin. SDS-PAGE indicated that decorin was present in all regions and fibromodulin was mainly observed in the tensile region. These results indicate that an external condition, in this case the area available for locomotion, might influence the synthesis of extracellular matrix components and the mechanical properties of the tendon.

Biomechanical and biochemical properties of chicken calcaneal tendon under effect of age and nonforced active exercise

This study investigated if nonforced active exercise alters the biomechanical and biochemical properties of calcaneal tendon during maturation. Chickens at 1, 5, and 8 months old were divided into two groups: caged and penned. Intact tendons were used for biomechanical analysis, but they were divided into tensile and compressive regions for quantification of hydroxyproline and glycosaminoglycans. The exercise increased tendon strength after the fifth month, energy absorption in the eighth month, and ultimate tensile stress in the first month. Age increased tendon strength and energy storage and reduced stiffness but did not alter stress. There was an increase in collagen content in the fifth month. Glycosaminoglycans showed a progressive decline in the tensile region. Thus, some biomechanical and biochemical changes depend on the maturation process itself and also are influenced by spontaneous exercise, showing that mechanical stimulation of low intensity may help to improve the quality of the tendon.

Arrabidaea chica extract improves gait recovery and changes collagen content during healing of the Achilles tendon

INTRODUCTION Tendon lesions are still a serious clinical problem. The leaves of the Bignoniaceae Arrabidaea chica (Humb. & Bonpl.) B. Verlot. (syn. Bignonia chica (Bonpl.)) have been used in traditional medicine and described in the literature for its healing properties. However, no study has shown the effects of A. chica during tendon healing. The aim of this study was to investigate the healing properties of the A. chica leaves extract on tendons after partial transection. METHODS A partial transection in the tension region of the Achilles tendon of rats was performed with subsequent posterior topical application of A. chica extract (2.13g/mL in 0.85% saline solution) at the site of the injury. The animals (n=154) were separated into 7 groups: N - rats with tendons without transection; S7, S14 and S21 - rats with tendons treated with topical applications of saline for 7 days and sacrificed on the 7th, 14th and 21st days after surgery, respectively; A7, A14 and A21 - rats with tendons treated with topical applications of the plant extract. The transected regions of the tendons were analyzed through biochemical, morphological and functional analyses. To evaluate the type and concentration of collagen, Western blotting for collagen types I and III was performed, and the hydroxyproline concentration was determined. The participation of metalloproteinases (MMP)-2 and -9 during tendon remodelling was investigated through zymography. Gait recovery was analyzed using the catwalk system. The organization of the extracellular matrix and morphometry were detected in sections stained with haematoxylin-eosin. RESULTS The application of A. chica extract in the region of tendon injury led to an increase in the amount of hydroxyproline (mg/g tissue) on the 7th (91.5±18.9) and 21st (95.8±11.9) days after the tendon lesion relative to the control groups treated with saline (S7: 75.2±7.2; and S21: 71.9±7.9). There were decreases in collagen types I and III (as determined by densitometry) in the groups treated with the plant extract 7 days after injury (type I: 103.9±15.9; type III: 206.3±8.1) compared to the saline-treated groups (type I: 165.2±31.1; type III: 338.6±48.8). The plant extract stimulated the synthesis of MMP-2 on the 21st day after the lesion and decreased the amount of latent and active isoforms of MMP-9 on the 14th day. Analysis by the catwalk system (max contact intensity) showed that the A. chica extract improved the gait of rats on the 7th day of the healing process when compared to the saline group. CONCLUSIONS The use of A. chica extract during the healing process of the tendon leads to an increase in collagen content and improved gait recovery. Further studies will be performed to analyze the effect of this plant extract on the organization of the collagen bundles of tendons after lesions and to study its probable anti-inflammatory effect.

Structural and Biochemical Analysis of the Effect of Immobilization Followed by Stretching on the Calcaneal Tendon of Rats

Little is known about the stretching effects on the biochemical and morphological features of tendons submitted to a long period of immobilization. Our purpose was to evaluate the response of rat tendons to stretching procedures after immobilization. The animals were separated into five experimental groups: GI—control of immobilized and euthanized animals; GII—immobilized and euthanized animals; GIII—control of immobilized animals and afterward stretched or allowed free cage activity; GIV—immobilized and stretched animals; and GV—immobilized and allowed free cage activity. Analysis in SDS-PAGE showed no remarkable differences among the groups, but a prominent collagen band was observed in GV, as compared to GIV and the control group, both in the compression and tension regions. Hydroxyproline content was highest in the compression region of GII. No differences among the groups were observed in the tension region. In regard to the concentration of noncollagenous proteins, differences were detected only in the tension region, where larger concentrations were found in the GII. When GII and GIV were compared, highest values were found in the GII. A more abundant presence of sulfated glycosaminoglycans, especially chondroitin sulfate, was detected in GIV, at the compression region of tendons. The presence of dermatan sulfate was outstanding in the compression and tension regions of the GII and GV groups. In the Ponceau SS stained sections, analyzed under polarization microscopy, GII exhibited the highest disorganization of the collagen bundles, partially recovered after stretching or with only remobilization. Our results indicate that a revision in the stretching procedures, in terms of duration and periodicity of the sessions, could benefit the efficiency of the stretching in cases of previous immobilization of tendons.

The effect of age on the structure and composition of rat tendon fibrocartilage

Biochemical and morphological aspects of fibrocartilages of calcaneal and deep digital flexor tendons in rats aged 30, 180 and 730 days were analyzed. In both tendons a stronger staining with Alcian blue, indicating the presence of proteoglycans, was detected in rats of 30 and 180 days. In animals 730 days old, it was restricted to the pericellular area. Ultrastructural analysis showed a more prominent pericellular matrix in calcaneal tendon compared to the deep digital flexor tendon. The biochemical analysis showed higher levels of proteins and glycosaminoglycans in the calcaneal tendon of 30‐day‐old rats compared to older rats. In the deep digital flexor tendon, no significant differences were observed between ages. The small proteoglycan, fibromodulin, was detected in both tendons of all ages, but in young rats it appeared to be running as a 210 kDa component, probably due to the association with collagen chains or self‐association.

Stroma-mediated granulocyte-macrophage colony-stimulating factor (GM-CSF) control of myelopoiesis: spatial organisation of intercellular interactions

Granulocyte-macrophage colony-stimulating factor (GM-CSF) is one of the major cytokines involved in control of haemopoiesis both in bone marrow and in extramedullar sites. Its biological activity depends upon the composition and physicochemical properties of the microenvironment provided by the supporting stroma. GM-CSF activity is modulated and controlled by the stromal heparan-sulphate proteoglycans, but their optimal interaction occurs only at low pH. We questioned whether the microenvironment organisation of the interface between stroma and haemopoietic cells provides such conditions. We studied myeloid progenitor proliferation in contact with bone marrow-derived and extramedullar stromas using electron microscopy and selective labelling of pericellular components. We present evidence that, upon interaction, the two cell types reorganise their interface both in shape and molecular composition. Haemopoietic cells extend projections that considerably increase the area of intercellular contact, and stromal cells form lamellipodia and carry out a redistribution of membrane-associated sialylated glycoconjugates and proteoglycans. Such rearrangements lead to extensive capping of negatively charged molecules at the interface between the supporting stroma and the haemopoietic cells, leading potentially to a local decrease in pH. Our results indicate that the distribution of negative charges at the cellular interface may be responsible for the selectivity of cell response to GM-CSF.

Effect of the Arrabidaea chica extract on collagen fiber organization during healing of partially transected tendon

AIMS After undergoing lesions, tendons have disorganized collagen fibers compared to undamaged tendons. Arrabidaea chica leaves have the aglycones carajurin and carajurone, components of the antocyanins, with a strong pharmacological potential due to their healing properties. Thus, the aim of this study was to investigate the effect of topical application of A. chica extract during tendon healing. MAIN METHODS The calcaneal tendon of Wistar rats was partially transected with subsequent treatment with A. chica extract (2.13 g/mL) followed by excision on the 7th, 14th and 21st days. Control rats received only saline treatment. KEY FINDINGS Transmission electron microscopy analysis showed the presence of a large amount of small segments of collagen fibrils in the transected region of the tendons on the 7th day in both the control and plant-treated groups. Considering the organization of the collagen fibers, higher values of birefringence were observed under polarization microscopy in the tendons of the plant-treated group on the 14th day compared to the control group. A larger quantity of dermatan sulfate was also detected after plant treatment in the same period. However, lesser dermatan and chondroitin sulfate were detected in the plant-treated group than in the control group on the 21st day. No differences were found in the values of birefringence between these groups. Intense metachromasy was observed in both transected groups on the 21st day. SIGNIFICANCE In conclusion, the use of A. chica extract improves collagen organization and increases the quantity of dermatan sulfate on the 14th day of the tendon healing.

Effects of passive stretching on the biochemical and biomechanical properties of calcaneal tendon of rats.

The role of physical activity in affecting the composition of extracellular matrix and mechanical properties of tendons has been well studied, but little is known about the role of passive stretching. The purpose of this study was to test the hypothesis that stimulation by passive stretching may change the composition and mechanical properties of tendons. Three-month-old Wistar rats were divided into three groups: the control, animals were not submitted to stretching procedures; groups that had their calcaneal tendons manually stretched three or five times a week, for 21 days. Afterward, the calcaneal tendons were removed and assayed for hydroxyproline content and biomechanical test. The hydroxyproline content in the stretched groups was higher, suggesting that more collagen was present in the tendons of these groups. These tendons also showed higher values of maximum stress and modulus of elasticity or Youngs modulus. These results indicate that stretching leads to alterations in the synthesis of the extracellular matrix components and in the mechanical properties of tendons.