Therapeutic Effects Of Quercetin On Rat Experimental Periodontitis

Objectives:To build a rat pericoronitis animal model through ligation of maxillary first molar. After intragastric treatment with quercetin for 6 weeks, the morphological changes, and expressions of cycloxygenase(COX-2) and matrix metalloproteinase(MMP-8) in periodontal tissues were investigated. The objectives are to investigate the therapeutic effects of quercetin on rat experimental pericoronitis and provide a theoretical basis and experimental reference for application of quercetin into treatment of pericoronitis in clinic.Methods:Totally 50 six-month-old healthy female Sprague-Dawley(SD) rats(each weight 300 g) were selected. The rats were adaptively fed for 1 week with foods and water. Then 10 rats were randomly selected and included into a normal group, followed by 6 weeks of normal treatment. After that, under anesthesia from intraperitoneal injection of 10% chloral hydrate, the rats were killed via femoral artery bleeding. Maxillary specimens were collected for analysis. Under anesthesia with 10% chloral hydrate, the other 40 rats were treated with ligation of bilateral maxillary first molars on cervix with use of ligature wire(diameter 0.20 mm). After daily feeding with high-glucose high-viscosity foods for 6 weeks, 10 rats were randomly selected and killed under anesthesia with intraperitoneal injection of 10% chloral hydrate. The maxillary specimens were collected for determination of pericoronitis modeling. These 10 rats were included as a ligation group. The other 30 rats were randomly divided into 3 groups(each n=10). The Removal Ligation group( R group) was treated with removal of ligature wires under anesthesia via injection of 10% chloral hydrate. Then after 6 weeks of normal feeding, the rats were killed and maxillary specimens were collected. The Removal ligation+ Quercetin group(RQ group) was treated with removal of ligature wires under anesthesia via injection of 10% chloral hydrate. Since the 7th week, each rat was treated with 100 mg/(kg.d) quercetin for 6 weeks. After that, the rats were killed and maxillary specimens were collected. The With ligation+ Quercetin group(WQ group) was treated with 100 mg/(kg.d) quercetin since the 7th week. After 6 weeks, the rats were killed and maxillary specimens were collected. All specimens were fixated in a 10% formaldehyde solution for 48 h and decalcified in 10% EDTA solution for 8 weeks. Then periodontal tissues continuous slices were prepared. The morphological changes of periodontal tissues were observed under a HE staining light microscope. The expressions of COX-2 and MMP-8 in periodontal tissues were detected with immunohistochemical staining. The results were sent into statistical analysis.Results: 1 Naked eye observationN group: The shapes of gingival papilla are basically complete, without swelling or bleeding. Periodontal pockets were not detected.L group: Gums are swollen and separated from the tooth face. Food residue accumulated there. Deep periodontal pockets were detected. Some periodontal pockets were found with secretion of purulent exudate and bleeding.R group: Gums are not swollen or bleeding. The shapes of gingival papilla are basically complete.RQ group: Gums are not swollen or bleeding. The shapes of gingival papilla are basically complete. This group is not significantly different from N group.WQ group: Gums are not swollen or bleeding. Periodontal pockets are shallow compared with L group. 2 Pathological observationN group: Junctional epithelia are located at the enamelo-cemental junction. No inflammatory cell infiltration occurred. Periodontal fibers are arranged regularly. No obvious absorption was found in the alveolar ridge crest.L group: Junctional epithelia are separated from the enamelo-cemental junction and displaced to the root part to form deep periodontal pockets. Abundant inflammatory cell infiltration occurred. The periodontal membranes were found with broadened gaps. Periodontal fibers are arranged irregularly and the surrounding blood capillaries were expanded. Active osteoclast bone absorption sinks appeared at the alveolar bones. Bone absorption disorders occurred at alveolar ridge crest and proper alveolar bone.R group: Periodontal pockets are located at the enamelo-cemental junction. Coarse large hyperplasia of collagenous fiber bundles appeared in between periodontal pockets and alveolar bones. Slight chronic inflammatory cell infiltration was found. New osteoid formation was found in the original bone absorption sinks.RQ group: Periodontal pockets are located at the enamelo-cemental junction. Nearly no inflammatory cells were found in the wall epithelia or junctional epithelia in the periodontal pockets. Small amounts of bone absorption defects and osteoclasts, but large amounts of osteoblasts were found in the alveolar bones. Osteoblasts were also found in alveolar ridge crest.WQ group: Periodontal pockets became shallow. Slight inflammatory cell infiltration occurred in wall epitheliaa and junctional epithelia. Abundant newly-formed fiber connective tissues and newly-formed blood capillaries were found in between periodontal pockets and alveolar bones. New bone formation occurred in alveolar ridge crest. 3 Distributions and expressions of COX-2 and MMP-8 in periodontal tissues 3.1 Expressions of COX-2 in periodontal tissuesIn periodontal tissues, COX-2 is mainly expressed in the infiltration zone with accumulation of inflammatory cells, and in vescular endothelial cells and fibroblasts of pavement epithelia and lamina propria. COX-2 positive staining shows brown yellow or yellow grains. The average optical densities are: N group 0.149±0.0042; L group 0.181±0.0050; R group: 0.165±0.0040; RQ group: 0.152±0.0021; WQ group: 0.162±0.0039. Results of statistical analysis are listed below. 3.1.1 COX-2 expression in L group is strongly positive(+++) and significantly different from N group(P<0.05). 3.1.2 COX-2 expression in R group is positive(++) or weakly positive(+) and significantly different from N group(P<0.05) and L group(P<0.05). 3.1.3 COX-2 expression in RQ group is weakly positive(+) or negative(-) and insignificantly different from N group(P>0.05), but significantly different from R group(P<0.05). 3.1.4 COX-2 expression in WQ group is weakly positive(+) and insignificantly from L group(P <0.05) and RQ group(P<0.05). 3.2 Expressions of MMP-8 in periodontal tissuesIn periodontal tissues, MMP-8 is mainly expressed in the infiltration zone with accumulation of inflammatory cells and in fibroblasts and osteoclasts. COX-2 positive staining shows brown yellow or yellow grains. The average optical densities are: N group 0.154± 0.0032; L group 0.203± 0.0057; R group: 0.169± 0.0022; RQ group: 0.158± 0.0022; WQ group: 0.162± 0.0046. Results of statistical analysis are listed below. 3.2.1 MMP-8 expression in L group is strongly positive(+++) and significantly different from N group(P<0.05). 3.2.2 MMP-8 expression in R group is positive(++) or weakly positive(+) and significantly different from N group(P<0.05) and L group(P<0.05). 3.2.3 MMP-8 expression in RQ group is weakly positive(+) and insignificantly different from N group(P>0.05), but significantly different from R group(P<0.05). 3.2.4 MMP-8 expression in WQ group is weakly positive(+) and significantly from L group(P <0.05) and RQ group(P<0.05).Conclusions:1 A rat experimental pericoronitis model was built through wire ligation of maxillary first molar and feeding with high-glucose high-viscosity foods.2 Treatment with pure quercetin can reduce the expression levels of COX-2,MMP-8 in periodontal tissues.3 Treatment with quercetin accompanied with removal of periodontal local stimulating factors can significantly reduce the expression levels of COX-2,MMP-8 in periodontal tissues.4 Treatment with quercetin accompanied with removal of periodontal local stimulating factors has significantly therapeutic effects on experimental periodontitis.