Effect Of Local Transplantation Of Gingival Mesenchymal Stem Cells On Experimental Periodontitis In Rats

Objective:Gingival mesenchymal stem cells are a type of stem cells with self-renewal,multi-directional differentiation,anti-inflammatory and immunomodulatory effects.Studies have shown that it is easy to separate,It also has a single morphology and a stable phenotype in long-term culture,and can maintain normal karyotype and telomerase activity.On the other hand,It has low immunogenicity and without tumorigenic.According to reports,it has the ability of osteogenic differentiation and has potential value in bone tissue reconstruction and regeneration.In this experiment,human gingival mesenchymal stem cells(GMSCs)were injected locally into the experimental rat periodontitis model constructed to observe its therapeutic effect on experimental periodontitis.Methods:1. Forty male SD rats were randomly divided into four groups:1)normal group(group N);2)saline group(group Na Cl);3)periodontitis group(group P);4)cell injection group(group C).Neck of bilateral maxillary molars of all rats in the group Na Cl,group P and group C were ligated with 0.2mm orthodontic ligature,and experimental periodontitis model was established with glucose-dipped water diet.Group N did not do any processing.2. Eight weeks later,one rat was randomly selected from each of thegroup Na Cl,the group P,and the group C,and the rats were sacrificed,and bilateral maxillary specimens were retained.The general inspection and pathological section images were used to verify whether the periodontitis model was successfully established.Then carefully and slowly removed the orthodontic ligature of the teeth of the other rats in the three groups.In group C,15ul of human GMSCs(cell density is 10~6/100ml,human GMSCs+Na Cl suspension preparation for injection provided by Stem Cell Laboratory of the First Hospital of Hebei Medical University)were injected into the mesial,central and distal of the gingival sulcus of the second molars.The injection depth is appropriate when the needle tip touches the bone surface and encounters a little resistance.Rats in the group Na Cl were injected with the same amount of saline at the same site.Injected once every two weeks,and two injections in total.Group P and Group N did not receive injections.3. After the fourth week,the femoral artery was drawn under abdominal anesthesia and the rats were sacrificed.The bilateral maxilla was immediately separated,and the right maxilla was fixed with 4%paraformaldehyde for 48hours.After 4 weeks of decalcification,HE staining analysis was performed.The data obtained by measuring the distance from the enamel cementum to the alveolar crest after mechanical separation of the bone from the soft tissue was recorded as the alveolar bone loss(ABL).Enzyme-linked immunosorbent assay(ELISA)was used to detect the levels of interleukin 1?(IL-1?)and tumor necrosis factor?(TNF-?)in rat serum.Results:1.Rats were sacrificed after 8 weeks of ligation.The grossspecimens were observed with the naked eye.The gingival redness and swelling of the ligament site of bilateral maxillary second molars were found.The calculus and plaque was accumulated around the neck.The periodontal pocket was formationed,and the alveolar bone crest was absorbed obviously,proving that the periodontitis model was successfully constructed.2.Compared with the group Na Cl,the ABL value of the group C was significantly reduced,and the difference was statistically significant(P<0.05).The ABL value of the group C and the group N was not statistically significant(P>0.05).3.The levels of IL-1?and TNF-?in group C were significantly lower than those in group Na Cl(P<0.05),and the difference was statistically significant.4.HE staining results showed that the gingival papilla of group P and group Na Cl were damaged,and the tissue was incomplete.There were a large number of inflammatory cells in the gingival tissue.The combined epithelium migrated to the root tip obviously,and the height of the alveolar bone crest was significantly reduced.For group C,there were a small amount of inflammatory cells was infiltrated in the gingival tissue,combined with the mild migration of epithelium to the root tip,and the alveolar bone crest was slightly absorbed.The gingival papilla of group N remained intact,the combined epithelium was attached to the enamel cementum and without the absorption of the crest bone crest,and also there was no obvious infiltration of inflammatory cells in the gingival tissue.Conclusions:1.The rat model of periodontitis was successfully established by orthodontic ligature ligation of the tooth neck and combined with glucose water diet.2.The local injection of human GMSCs has an anti-inflammatory effects in rats,reducing the expression of inflammatory factors IL-1?and TNF-?in the serum,inhibiting the bone resorption and producing a certain therapeutic effect on periodontitis.