Experimental Study On Vital Pulpotomy And Repair Of Dental Pulp After Nd: YAG Laser Irradiation

Objective:Through establishment of animal model of vital pulpotomy with dogs irradiated by pulsed Nd:YAGlaser at the dose of certain energy parameters,to observe the expression of HSP70, TGF-β₁,BMP-2 at different periods and regions in dog's dental pulp (DP) and the microscopic changes under transmission electron microscope(TEM). The biological significance of these changes during the injury defense and self-reparative process are analyzed to provide insight into the injury and repair mechanism of dental pulp after laser irradiation.Methods:1. Four healthy local mongrel dogs were selected and the teeth were prepared for vital pulpotomy,each tooth was irradiated by Nd:YAG laser with three different parameters. And the one of the four dogs was killed after 3 days and the teeth were extracted, immediately fixed in 4% formaldehyde-0.1 mol∕L PBS solution. The other three dogs were killed respectively at 15,21 and 45 days after lased and the teeth were extracted and treated as above.2. Four healthy local mongrel dogs were selected and the teeth were prepared for vital pulpotomy, were made into models of vital pulpotomyand fed routinely and were randomly divided into namber1-4.each: 1 (sacrificed after 3 days of laser irradiation); 2 (sacrificed after 15 days of laser irradiation); 3 (sacrificed after 21 days of laser irradiation); 4 (sacrificed after 45 days of laser irradiation); To employ 96 specimens from dogs as experiment teeth. 4 Control group were labeled as group A₁ , A₂,A₃ and A₄. Each group has 12 premolares and molares taken from left hand side upper jaw and lower jaw of one dog. 4 Study groups were labeled as B C D E respectively. Each group has 12 premolares and molares taken from the right-hand side upper jaw and lower jaw of one dog. After sacrificing the dogs, 80 specimens (10 for each group), which were taken from dogs'upper right premolares and molares combined with some jaw bones, were acquired immediately for HE staining and immunohistochemical analysis. 2 premolares of a dog randomly from each group was split and the freshly pulp was obtained for TEM observation. Average optical density (OD) values of all immunohistochemical images and results of HE staining were measured with Image-Pro Plus 5.0 image analysis software. Ultramicrostructural changes of DP were observed and analyzed. Statistical methods: Kruskal-Wallis method, Nemenyi method, one-way ANOVA and LSD-t test with SPSS 13.0.Results:1. The subsequent histological examination of the irradiated teeth showed that the wound healing of the irradiated pulp seemed to be better than that of controls at 3 days, Dentin bridge formation in the irradiation pulp was facilitated at 15,21 and 45 days after operation.5 s irradiation at 40 Hz, 50 mJ (2 W) promoted the pulp wound healing and complete hard tissue bridges formation with less pulpal inflammation in all the dogs in this group. 5 s irradiation at 60 Hz, 50 mJ,(3 W) caused unreversible pulp damage.2. there were statistical difference between B and A,B and C, B and D during 3 days after lased(P<0.05), there were statistical difference between B and C, B and D during 15,21 or 45 days after lased (P<0.05). statistical results of vacuole change of the odontoblast and vascular engorgement, hemorrhage of dental pulp were as following: there were statistical difference between B and A₁, B and A₃, B and A₄, B and D, B and E (P<0.05). Inflammation cell infiltration of DP came to peak in Group C and recovered normal after 45 days. Some tertiary dentine was observed in Group E.Discovery under TEM: Group B and Group C: Perinuclear space of odontoblasts distended slightly; rough endoplasmic reticulums (RERs) distended mediumly, some of them were vesicle shape and presented degranulation phenomena; a part of cristae mitochondriales (Mi) and matrix were degraded. Phagocytic cells'bodies were large with multi-ecphyma, plenty of cellular organs, lysosomes and phagocytotic vesicles. Group D and Group E: similar to Group A_1-4, there were plenty of cytoplasm, RERs, ribosomes (RIs), mitochondrias, well-developed Golgi's complexes which have multilayer applan vesic, bu and bullule. RER had a lot of attached RI and free polyribosomes. Mi was orbicular-ovate and has cristae lamellar.3. Statistical results of immunohistochemistry staining: HSP70 expression state in coronal pulp: Group B has statistical differences with the other groups. TGF-β₁ expression state in coronal pulp: Average OD value of Group D was highest in 8groups. BMP-2 expression state in coronal pulp: Average OD value of group E was highest in 8 groups.Conclusion:â‘ DP undergone reversible inflammation changes after laser irradiation at the dose of 40 Hz,2 w,50 mJ which can be used safely. Within high energy parameters, pulsed Nd∶YAG laser may cause unreversible pulp damage. The extent of the pulpal injury seemes to depend more on the exposure time than on the output power.â‘¡Ultramicro structural changes showed that DP had the ability of self defense and repair.â‘¢HSP70 arose specially when DP received heat stimulation and played a role during the later period of pulp repair.â‘£TGF-β₁ and BMP-2 were presented in normal or impaired DP. The dynamic changes of expression showed that the two factors defended DP in the early period, then participated in the self-reparative process and promoted the formation of reparative dentin..