| Objective: As an important part of the temporal-mandibular joint, the bone remodeling process and stability of the mandibular condylar is the key for the research. The growth of the condylar is highly related with the external mechanical stimulus. After the condylar cartilage cells are stimulated by external relevant factors(especially the mechanical stress) and the corresponding signal transduction is passed, they will display a series of biological responses such as cell proliferation, differentiation and programmed death. However, the way how the relevant cell factor adjusts the cartilage creation and endochondral ossification and completes the bone remodeling needs further research. DMP-1 is a kind of non-collagenous protein belonging to small integrin-binding ligand N-linked glycoprotein(SIBLINGs), as a mineralization factor, it participates in the mineralization of hard tissue and is separately expressed in cartilage cells and bone cells. It is an essential special mark for cartilages and bones. In recent years, it finds that it the adaptable expression appears in the DMP-1 in the bone cells after the mechanical stress stimulation. DMP-1’s participation as a transcription factor in the bone remodeling causes public attention, the formation, development and remodeling of the mandible is closely related with the expression of DMP-1 genes. Meanwhile, PCNA only exists in the proliferative cells and tumor cells and it is one of the marks for cell proliferation and it can effectively help reflect the proliferation function of cartilage cells.It adopted histological staining and immunohistochemical staining and other methods in this experiment. In different times, detected the expression of DMP-1 within the condylar cartilage during the process of mandibular functional protrusion of Sprague-Dawley rats, its change rules, and theexpression of PCNA, so as to study the function of DMP-1 on the cartilage proliferation in the condylar bole remodeling during the process of mandibular functional protrusion and provide experimental evidence for the clinical orthopedic treatment for patients with bone type II malocclusion.Methods: 70 healthy male Sprague-Dawleys(SD) rats in the age of 4 weeks and weight about 90 g were selected. After a week of adaptable feeding, randomly and equally divided the rats into experimental group and control group. According to the experimental period(1, 3, 7, 14, 21, 28 and 35days), respectively divided each group into 7 subgroups, 5 SD rats for each subgroup and 14 subgroups in total. These experimental groups would make use of 3M glass ion binding material to bond the self-made mandibular protrusion bite plate appliance on the upper incisors of rats. Selected rubber bands with appropriate models and connected the two sides of face bow over the nasomaxillary complex and made the bite plate fixed. Closely wrapped the limbs of rats with medical adhesive plasters to effectively prevent the damage of the forepaws of rats on the device and the injury to facial tissues. The appliance should be worn all day. Didn’t make any experimental operation on the control group. All animals shoud be fed with soft diets, drink water freely, and fed them under the same environment till the end of the experiment. The rats were taken out for conventional fixation, decalcification, dehydration and embedding on the 1, 3, 7, 14, 21, 28 and 35 days of the experiment. After the paraffin section, the routine HE staining observation on condylar histologic changes shoud be made; detected the expressions of DMP-1 and PCNA by immunohistochemical method, adopted the cell image analysis system to make the rating for immumohistochemical staining and took HIS value to show the staining intensity. All the data results shoud be processed by SPSS21.0 statistical software. When making the statistic analysis, adopted t test for the comparisons between the experimental group and control group, and adopted one-way ANOVA method for intra-group comparison. Bivariate correlation analysis was used to the experimental group DMP-1 and PCNA. the test level α=0.05.Results:1 General ObservationThe mandibles of SD rats in the experimental group were in the status of moderate protrusion, their upper and lower anterior teeth were edge to edge bite or slightly cross bite. After moving the appliance, the mandibular could not retreat, while overjet of anterior teeth of the SD rats in the control group was about 3mm.2 Histological FeaturesThe mandibular condylar had a unique domain layer, which couid be roughly divided into four layers from the surface to the deep structure: fibrous articular layer, cellular rich zone, fibrocartilaginous zone and zone of calcified cartilage. For the control group, a layer of fibrous tissue(the intrarticular disc) could be seen wrapping around the surface of the condylar cartilage under the microscope, the front of the condylar cartilage was the thinnest and the rear part was the thickest. The condylar cartilage was subject to age-related changes over time, the changes in central and rear parts were more obvious, the changes in fiber layer and cellular rich zone were not obvious, while the thinning of fibrocartilaginous zone was more obvious. For the experimental group, under the same point in time, the thickness of condylar cartilage of SD rats was increased compared with the control group, especially at the rear of the condylar cartilage, while the variation in thickness of the front part was not obvious and the pathological changes were not found. The cellular layers of cellular rich zone and fibrocartilaginous zone were increased, the cell volume of the hypertrophic chondrocytes was enlarged and the extracellular matrix was increased.3 Immunohistochemical ResultsThe negative control group(PBS insteads of primary antibodies) showed no positive staining. In the condylar cartilage cells of the control group, DMP- 1 mainly expressed in fibrous cartilage layer and maintains a relatively low level, higher from the 3rd to the 14 th day, comparing differences between groups had no statistical significance(P>0.05). In the 3rd day for the experimental group, the DMP-1 protein expression in condylar cartilage was obviously enhanced, IHS=44.67±3.05, which had significant difference(P<0.01) with control group IHS=10.33±0.58; on the 7th day, the experimental group IHS=50.67±7.02, the positive expression was increasingly strengthened, which had significant difference(P<0.01)compared with control group IHS=10.67±1.15; on the 14 th day, the experimental group IHS= 85.67 ± 4.51, the expression of DMP-1 reached to the highest level, which had significant difference(P<0.01) compared with control group IHS=10.67±1.53 and the 7th day of the experimental group; on the 21 th day, the experimental group IHS=42.67±10.07, the positive expression cells of DMP-1 began to decrease, but they had significant difference(P<0.01) compared with control group IHS=10.00±1.00; Until the 28 th day, they had significant difference(P<0.01) for the experimental group IHS=16.67±1.15 compared with control group IHS=10.00±1.00; on the 35 th day, the protein expression DMP-1 of condylar cartilage fell back to a lower level, the experimental group IHS=10.00±1.00, there were significant differences(P>0.05) compared with the control group IHS=9.67±0.58.PCNA was located at condylar cartilage cellular rich zone and shallow hypertrophy cartilage cell layer, the PCNA expression in condylar cartilage cells began to increase since the 3rd day of the experimental group, IHS=49.67±13.50, which had significant difference(P<0.01) compared with control group IHS=7.33±1.15; on the 7thday of the experimental group IHS=124.0±7.55, which had significant difference(P<0.01)compared with the control group IHS=9.00±2.00; on the 14 th day, PCNA positive expression of experimental group began to decrease, IHS=108.33±8.50, but they had still significant difference(P<0.01) compared with the control group IHS=8.67±0.58; there were no statistical significances(P>0.05) for the experimental group and control group on the 21 th, 28 th and 35 th day.By bivariate correlation analysis, the resulted showed that the correlation coefficient of experimental group DMP-1 and PCNA was 0.825, P<0.01, their differences had statistical significance.Conclusions: 1 DMP-1 participated in condylar cartilage adaptive remodeling during the process of mandibular protrusion of rats, and promoted condylar cartilage proliferation and endochondral ossification.2 DMP-1 possibly participated in the process of transduction process from mechanical stress to biological signal during the bone remodeling of condylar cartilage of rats. |
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