动物造模 z-bio 2021-11-25 714

　　Background: Anterior cruciate ligament (ACL) has been increasing in frequency in reports of injuries to athletes with immature bones, accounting for 31% of knee injuries. Youth ACL reconstruction is still under discussion. Due to the risk of epiphyseal injury, non-surgical treatment is an attractive option. However, recent literature consistently shows that conservative treatment of ACL tears in children leads to a higher rate of instability and may progress to intra-articular injuries, including meniscal tears. Carry out early surgical treatment to restore immature kneecap stability. Although tendons have different biological and histological characteristics compared with ligaments, many studies have reported the application of tendons to repair ligaments. One of the still controversial issues is to clarify how this so-called "new bond" really gradually loses its tendon and ligament histological characteristics and exhibits specific biological characteristics. Although there has been a lot of research exploring various aspects of adult ACL reconstruction. However, there are few data on the tendon healing process in human or animal bone immature patients. Some animal studies have shown that implanted tendons do indeed seem to be remodeled into ligament ACL structures. The reconstruction of the anterior cruciate ligament in children requires the preservation of the epiphysis. Surgery should respect the biology and biomechanics of anterior cruciate ligament reconstruction, and minimize the possibility of affecting the growth of the lower limbs. Different tendons and bones have been proposed for preoperative cruciate ligament reconstruction in patients with immature bones, but the most commonly used is semitendinosus. The purpose of this study was to evaluate the morphological and histological changes in the articular cavity of a semitendinosus tendon to reconstruct the anterior cruciate ligament in rabbits and maintain its distal insertion. This study was conducted on growing animals to determine the type and extent of lesions. Immature animals have greater potential for tissue regeneration.

　　Method: This experimental study is a qualitative description of the ultrastructural changes of semitendinosus muscle used in anterior cruciate ligament reconstruction in young rabbits. The experiment was approved by the local ethics committee and followed our institution's animal care and use guidelines. Preoperatively, the animals received the antibiotic amoxicillin (10 mg/kg), two additional doses were given at 72-hour intervals, and 30 mg of ketorolac analgesia was continued for 2 days after the operation. All healthy rabbits entered the study. The exclusion criteria were claudication, pain, or other signs of disease such as septic arthritis. 21 New Zealand white rabbits, weighing about 1500±200 g, were operated on the right knee, and the left knee was used for comparison. The animals were anesthetized with diazepam (3 to 5 mg/kg) and ketamine (500 mg/kg). Expose the right knee joint through the medial parapatellar incision. The ACL is accurately removed, and the original footprint is marked with a sterile pencil. An oblique 2 mm diameter tunnel was drilled through the growth plate through the initial attachment of the ligament on the mid-tibia and dorsal femur. The semitendinosus tendon is severed at the junction of its proximal muscle and tendon, but the left Jian is attached to its distal end. The tendon passes through the tibia and femur tunnel, and then sutures the periosteum on the outer side of the injured femoral metaphysis with No. 2 silk thread. After the operation, it was fixed with plaster for seven days. Make every effort to reduce pain and avoid burdens. After the operation, the animals returned to their cages, free to eat and drink. The animals were randomly divided into seven groups, with three animals in each group, and were sacrificed at 1, 4, 6, 8, 12, 24, and 48 weeks after surgery. The right hind limb of each animal was dissected, and the deformity and unequal length of limbs were detected with calipers and radiography. The left knee was used to compare the normal ultrastructure of ACL. The specimens of the experimental group and the control group were fixed with 10% formalin. Then use EDTA for decalcification and paraffin embedding. 5μm thick sections were stained with H&E, Masson trichrome, Alcian-PAS, etc. Evaluate the sample under light microscope to analyze the changes in the joint cavity of the graft. All specimens were compared with normal rabbit anterior cruciate ligament structure.

　　Results: Any complications related to animals were observed throughout the study period. All grafts were intact at the time of evaluation, without the pearly appearance of a normal ACL. No failed semitendinosus grafts were observed at the time of the animal's death. One week after surgery, the transplanted tissue showed focal acellular necrosis. It can better evaluate tissue necrosis, such as a strong inflammatory reaction, after 3-4 weeks. But it was no longer detected within 40 days after the operation. A large number of fibroblasts appeared one month after the operation, and a large number of collagen fibers were arranged disorderly. Fibrous collagen, such as Type I and III, provides the basic building blocks of tendons and ligaments, with elasticity and strength. Fibroblasts are initially arranged longitudinally in the center of the tendon. The blood vessels are still poor, and the blood vessels are located around the graft. In the semitendinosus tendon, the fibroblasts are spindle-shaped, which is different from the anterior cruciate ligament fibroblasts. However, after 4 weeks, these spindle cells became more rounded. After the first month, the collagen fibers are arranged in disorder and the collagen fibers become more organized. In the fourth to eighth weeks after surgery, the typical inflammatory cells gradually disappeared. No necrotic area was detected. At 8 weeks after the operation, the collagen fibers arranged in parallel with the newly formed capillaries and became highly differentiated fibroblasts. Semitendinosus tendon, initially without blood vessels after transplantation, gradually becomes blood vessels, especially after 40 days. However, transplantation at this point in time did not appear similar to ACL. Compared with the normal left knee ligament, the average graft cross-sectional area of the operated knee joint is significantly higher. During the healing process of the new ligament (24 weeks), as the diameter of collagen fibers changes, the normal collagen direction and curling pattern disappear. From large to small, type I collagen fibers decrease and type III collagen synthesis increases. The histological characteristics of the animals sacrificed after 24 weeks did not change further within 48 weeks. Between 24 and 48 weeks, the graft has not yet acquired the characteristics of normal rabbit ACL in the articular cavity.

　　Conclusion: The immature rabbits did not form a ligamentation process for up to 48 weeks after surgery. The transplanted ST becomes a new entity during the healing process, with a microstructure between the tendon and ligament. The semitendinosus transplantation has undergone a process of adaptation, rather than completely restoring the biological characteristics of the complete ACL.