Effect Of Daidzein On The Healing Process Of Tooth Extraction And The Expression Of RANIKL In Ovariectomized Rats

Objective: To investigate the effects of phytoestrogen—Daidzein on thehealing process of tooth extraction and the expression of RANKL in ovari-ectomized rats.Materials and methods: Fortyfive female Sprague-Dawley rats (3monthsage)were submitted to ovariectomy(OVX) or sham surgery, and divided intothree groups: Sham, OVX and OVX/Da(ovariectomy with daidzeintreatment,10mg/kg.bw/day). The12-week treatment started8days afterovariectomy(the OVX/Da group were given a gavage of daidzein suspensions10mg/kg.bw/day,while Sham and OVX group were given distilled water withthe same way). The right mandible incisor were extracted at10days afterovariectomy to allow analysis of4,8and12weeks of alveolar healing process.After obtaining the histological samples, slides were stained with hematoxylinand eosin or subjected to immunohistochemical reaction for RANKL. Resultswere quantitatively evaluated by Image analysis software.Results:1. Hematoxylin and eosin staining histological examination: Atthe4weeks after tooth extraction, the Sham group and OVX/Da group showedmore newformed trabecular bone than the OVX group significantly, while theOVX group showed large empty area of blood clots, surrounded by fibroustissue, only few newformed trabecular bone was found.At8weekspost-extraction,the blood clot was completely absorbed in the OVX group,andthe trabecular bone number had increased, more fibrous connective tissue wasfound;the Sham group showed an established healing process, mature bonetrabecular and bone cell were observed,osteoblasts turned into bone-lining cells;the OVX/Da groupe showed thicker bone trabecular surrounded byfibrous connective tissue with active bone formation. At12weekspost-extraction,the thin trabecular bone of OVX group had interconnected, withlarge gaps fullfilled by fibrous connective tissue; the Sham group showedfurther bone remolding with bone resorption fossa;the OVX/Da group showedan established alveolar healing process with matured and regular trabecularbone.2. Bone histomorphometry analysis—Trabecular area ratio: the boneformation value of OVX group was significantly lower than the Sham groupand OVX/Da group in the post-extraction period(P<0.01).The Sham groupreached its maximum value at8postoperative weeks.The value of OVX/Dagroupwas significantly lower than the Sham group at4,8postoperativeweeks(P<0.01), but reached the same level with the Sham group(P>0.05) at12postoperative weeks.3. Immunohistochemical staining analysis:At4weeks after the toothextraction,the expression of RANKL was observed in fibrous connective tissuein each group,the gray value between each group was significantlydifferent(P<0.01).At8,12weeks after the tooth extraction, the new formedbone tissue did not express RANKL in both Sham and OVX/Da group,while inthe OVX group,bone cells in newformed trabecular bone expressed RANKL,and gradually increased, statistical analysis of positive cells showed significantdifference(P<0.01).Conclusions: Ovariectomy delays the alveolar healing process andinterferes bone formation through enhancing the expression of RANKL in bonetissue and cells.Low-dose daidzein treatment could promote the healing process,with an increase in the number of new-formed trabecular bone, probablythrough reducing the expression of RANKL in bone tissue, subsequently thedifferentiation and maturation of osteoclast was inhibitted.