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Study Of Autologous Platelet-rich Plasma On Rabbit Articular Cartilage ? Degree Damage Repair

Posted on:2017-01-29Degree:MasterType:Thesis
Country:ChinaCandidate:J ZhangFull Text:PDF
GTID:2334330488497947Subject:Surgery
Abstract/Summary:PDF Full Text Request
Objective:Cartilage injury is common in joint surgery disease, which without effective treatment mostly developed into osteoarthritis. In the later time, the change of joint biomechanics will be cause the pain, deformity and dysfunction, and it will be increase patients' psychological or mental burden. The traditional treatment methods of Medicine conservative treatment and cartilage transplantation or arthroscopic drilling decompression were not achieved satisfactory results. In recent years, along with the mechanism gradually in-depth research of autologous platelet-rich plasma (platelet-rich plasma, PRP), autologous platelet rich plasma also has certain curative effect in repair of cartilage injury in the Department of orthopedics. PRP has become a research hotspot, because a large number of various growth factors of PRP has a promoting effect on proliferation of chondrocyte and synthesis of extracellular matrix. But the research of PRP on II degree of articular cartilage injury is very poor. In this study, through the study of PRP on II degree of articular cartilage injury, to compare the degree of articular cartilage repair between the experimental group and control group, and provide a theoretical basis for clinical guidance.Methods:1.Grouping:Select 36 healthy New Zealand white rabbits as the experimental subjects, which were randomly divided into PRP, HA(hyaluronic acid) and NS(normal saline) three groups, each group of 12. Three groups of experimental animals were put to death at twelve weeks after surgery.2.Blood collection:using vacuum blood collection tube which contain sodium heparin anticoagulant to collect 8ml blood on ear central artery of each experimental animals, and using vacuum blood collection tube which contain EDTA-K2 to collect 2ml blood.3.Centrifugal: we apply twice centrifugation to prepare for PRP. After balancing centrifuge, it's the first time centrifugation of the speed 2000r/min, time 10 minutes. After the first time centrifugation, we absorbe the red blood cells under boundary layer 2mm and abandon. Puting the remaining part into the centrifuge and balancing centrifuge, it's the second time centrifugation of the speed 2000r/min, time 10 minutes. After centrifugation with a syringe from serum the layer above the 3/4 part and discarding the remainder, which is PRP. The 8ml rabbit arterial blood can be prepared about 0.8ml PRP.4.Platelet count:Take 2ml blood with EDTA-K2 anticoagulanted and PRP into hemocytometer to get the number of the platelets.5.Cartilage injury modeling and intervention:Each experimental animals are using the anterior median incision to open the skin and the medial parapatellar approach open knee joint capsule. Fully exposed rabbit knee medial condyle and manufactured II degree of cartilage damage model on the middle of the medial condyle positions by a Kirschner wire beam(an area of about 2 x 3mm2, a depth of about 0.3mm). Then, the PRP group left knee joint cavity inject 0.8ml PRP and 50ul activator(calcium gluconate, bovine thrombin), the HA group left knee joint cavity inject 0.8ml HA, and the NS group left knee inject 0.8ml saline. Experimental animals are not restrict activities after postoperative.6.Observations:After execution of experimental animals by air embolism method, open the left knee capsule and expose Cartilage injury for general observation. Apply the International Cartilage Repair Society (the International Cartilage Repair Society, ICRS) to evaluation. Then drawning from the repair site and making-up the paraffin sections, it was stained with HE, Safranin-0 and immunohistochemical staining. It was observed under a microscope and application of O1 Driscoll histological score to evaluated. IPP (Plus Image-Pro, IPP) 6.0 image analysis software was used for quantitative analysis of the gray value of immunohistochemistry of collagen type II collagen by immunohistochemical staining.Results:At 12 weeks after operation among the groups showed significant difference. In the PRP group, the surface of damaged area repaired better, smoother surface. In the HA group, there still can see the traces of damage. And in the NS group, the surface of damaged area repaired poor, rough, uneven. HE staining showed that the PRP group and normal newborn cartilage from cell morphology, cell distribution and the distribution of extracellular matrix are similar, and the chondrocyte proliferation is more. The HA group of new cartilage cells fewer than the PRP group. In the NS group, the number of newborn chondrocytes is the least, and there are significant differences in the morphology, cell distribution and extracellular matrix between the new and normal cartilage tissues. Safranin-O staining observed also showed that the damage area of the PRP group is similar to the normal area. There are large Safranin-O coloring cartilage matrix, and chondrocyte proliferation is strong in the HA group. The chondrocyte proliferation of NS group is fewer, and the mostly hyperplasia cells are fibroblasts. In immunohistochemistry, there has a large number of collagen type II collagen in injury region, and the damage area of the PRP group is similar to the normal area. The surface of HA group cartilage injury region lack of type II collagen, and cartilage subsurface shows a small amount of collagen type II collagen. In the NS group, it see injury area by a small amount of type II collagen and a large number of granulation tissue filled, obviously to see type II collagen immunohistochemical light stained area, damage repair area and normal tissue boundaries between more obvious, and the boundary is obvious between injury area and normal tissue.Statistical analysis:the three groups were analyzed by one factor analysis of variance (ANOVA), between every two groups were analyzed by least significant difference-t (LSD-t).The ANOVA results of general observation score results of three groups show statistically significant differences (F= 9.252, P< 0.05)), and the LSD-t results between each two groups show statistically significant difference(P<0.05).The ANOVA results of HE and safranin-O staining under the microscope observation of three groups show statistically significant differences (F=9.157, P< 0.05), and the LSD-t results between each two groups show statistically significant difference(P<0.05).The ANOVA results of mean density of Immunohistochemical staining of three groups show statistically significant differences (F=9.480, P< 0.05), and the LSD-t results between each two groups show statistically significant difference(P<0.05).Conclusion:This study confirmed:1. PRP for cartilage repair has a positive promoting effect. It can promote the repair speed after articular cartilage injury, and stimulates cell regeneration of cartilage matrix formation, to restore the original shape and structure, enhance the effect of the repair ofcartilage damage.2. PRP and HA have a repair effect on rabbit knee articular cartilage injury, but the effect of PRP on cartilage repair is better than that of HA.3. PRP have a better effect on II degree injury of rabbit knee articular cartilage, to provide guidance to the clinic.
Keywords/Search Tags:Platelet rich plasma, Cartilage repair, Chondrocyte, Cartilage injury, Cell proliferation
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