| Objective: To provide experimental and theoretical evidence to test if orthodontic functional treatment can be carried out on diabetics, by exploring the effect of diabetes mellitus on the remodeling of condylar cartilage during forward mandibular positioning under the treatment with insulin.Method: 1 One hundred and twenty male(3weeks of age) Sprague-Dawley rats (weight from 50 to 60g) were chosen and randomly divided into three groups with 40 rats each: Group A, the normal group; Group B, diabetes group and Group C, insulin treated group. Each group was randomly divided into five subgroups. 0, 7, 14, 21, 30 days group, consisted of 8 rats each. Streptozotocin (STZ) were intraperitoneal injected in rats of Group B and Group C with the amount of 85mg per kilogram to induct type 1 Diabetes mellitus. Rats in Group A were injected with the same amount of sodium citrate. Seventy-two hours later, blood was extracted from caudal vein and urine was sample before blood glucose and urinary glucose were respectively tested. Those rats whose blood glucose were above 16.8mmol/L and urinary glucose were three plus to four plus were diagnosed as diabetes. The fourth day after the model of diabetes mellitus was finished, rats in Group C began to receive subcutaneous injection of 0-4U insulin per day at their necks. The amount of injection was adjusted according to the rats'blood glucose and urinary glucose till the blood glucose was 8-10 mmol/L. Group A and Group B were given the same amount of normal saline injection. Ten days after the model of diabetes mellitus was established, all of the rats were equipped the appliance. Under the anaesthesia of 3% Nembutal, appliance was adhered on the upper incisor. The appliance was consisted of the upper crown and the inclined plane, and the inclined plane was made lingually by 25°. On adhering, the lower incisor was just on the labial inclined plane and the mandible was in an edge-to-edge or even cross-bite position when the rats bit, to guide the mandible on a forward position and molars untouched. During the experiment, all rats ate and drank freely. Animal feed was nutritional granula prepared by The Experimental Animal Center of HeBei Medical University. On adhering, all rats ate runny animal feed. The appliance must be used 24hours everyday. The rats were observed carefully and the new appliance was adhered in time if necessary.2 Testing of blood glucose: During the experiment, the blood glucose of Group B and Group C were measured every day, the rat whose blood glucose was not consistent with the experiment standard was rejected. Blood glucose have to be tested before rats were killed. Blood glucose of Group B was above 16.8 mmol/L. Blood glucose of Group C was less than 10 mmol/L and the rats of Group C were chosen to be examined for results.3 Preparation of the condylar cartilage histological sections: Group of qualified rats were killed by abstracting blood from their hearts respectively after adhered the appliance 0, 7, 14, 21 and 30 days. Took two-side condylar tissues (including articular disk, condylar cartilage and the osseous tissue under cartilage) of each rats, and then the specimens were infiltrated in 4% Polyoxymethylene 4°C for 72 hours. After decalcification in 7% EDTA 4°C about 6 weeks, anhydration and embedment, histological section were made by routine method. Then crosscuted the specimen through the median sagittal plane, and sliced the specimen continuously. Overviewed the condylar cartilage after functional treatment, by routing HE stain, collagen and mucopolysaccharide histochemical stain. Immunohistochemistry technique was used to detect the expression of TGF-β1 and VEGF in the condylar cartilage.Results:1 Histology: The thickness of the posterior part of condylar cartilage increased in groupA, groupB and groupC. There are no significant differences in the thickness of the cartilage between Group A and Group C. The area of the proliferative zone increased most obviously. The hypertrophic zone and the erosive zone also increased. The change in the resting zone was not obviously. The thickness changed firstly in the proliferative zone, and the change in two and three week was most obviously.The thickness of the anterior part of condylar cartilage decreased in groupA, groupB and groupC. There are no significant differences in the thickness of the cartilage between Group A and Group C. The thickness of Group B is thiner than that of GroupA and GroupC. The thickness changed firstly in the proliferative zone, but the change of the thickness concentrated in the hypertrophic zone and the erosive zone, the area decreased obviously. The change in the Resting zone was not obviously, and the change in three and four week was most obviously.2 Collagen: Every layer of the posterior part of condylar cartilage have bluish-green color in GroupA, GroupB and GroupC, and the color in the resting zone and the hypertrophic zone is heaviest. The color is taper from0day, 1 week to 2 week, there is no difference between 3 groups. The color of the proliferative zone, the hypertrophic zone and the erosive zone is light green in 3 week, there is no significant differences between GroupA and GroupC. The strength of color of GroupB attenuated more obviously in 3 week. The strength of color of the hypertrophic zone and the erosive zone of GroupA darkened in 4 week. The strength of color of the hypertrophic zone of GroupC darkened in 4 week too. Every layer of GroupB has not changed obviously in 4 week.3 Acid mucopolysaccharide: The coloration of the acidic mucopolysaccharide of GroupA, GroupB and GroupC mainly resided in ground-substance, and more concentrated in the hypertrophic zone. The positive reaction winded up gradually from the proliferative zone to the erosive zone, bone trabeculah had positive reaction too. The coloration of the acidic mucopolysaccharide was taper from 0day, 1 week to 2 week. There was no difference between 3 groups. The coloration of the proliferative zone, the hypertrophic zone and the erosive zone of GroupA and GroupC thined and maldistributed. The blue-stain confine decreased in 3 week. There was no significant differences between GroupA and GroupC. The strength of color of GroupB was weaker than GroupA and GroupC. The strength of color of the hypertrophic zone and the proliferative zone of GroupA lightly deepened in 4 week. The coloretur of the hypertrophic zone of GroupC deepened in 4 week too. The coloretur of GroupB did not deepened in 4 week.4 TGF-β1: Every layer of the posterior condylar cartilage had immunostaining of TGF-β1 in GroupA, GroupB and GroupC. The highest expressions was in the hypertrophic zone. The immunostaining of GroupA gradually increased from 1 week, the masccline was strongest in 3 week and the backswing tendency appeared from 4 week. The immunostaining of GroupB gradually increased from 1 week, and the backswing tendency did not appear. But the strength of the immunostaining was lower than GroupA. Group B has significant differences with Group A (P< 0. 01). The immunostaining of GroupC gradually increased from 2 week, and the backswing tendency did not appear too. Group C has no significant differences with Group A (P> 0. 05).5 VEGF: The resting zone of GroupA, GroupB and GroupC did not express VEGF, and the expression in the proliferative zone was weak, the expression gradually increased from the proliferative zone to the upper 1/3 of the hypertrophic zone. The expression was strongest in the middle 1/3 of the hypertrophic zone, and the expression began to weak in the below 1/3, the expression was weakest in the erosive zone. The immunostaining of GroupA gradually increased from 2 week, the masccline was strongest in 3 week and the backswing tendency appeared from 4 week. The immunostaining of GroupB gradually increased from 2 week, and the backswing tendency did not appear. But the strength of the immunostaining was lower than GroupA, Group B has significant differences with Group A (P< 0. 05). The immunostaining of GroupC gradually increased from 3 week, and the backswing tendency did not appear too. Group C has no significant differences with Group A (P> 0. 05). The strength was lower than GroupA, but higher than GroupB . Group C has significant differences with Group A and GroupB (P< 0. 05).Conclusion: 1 During the experiment on the diabetic rats, quantity of the cell of condylar cartilage in the anterior part decreased obviously, and the renovation of cartilage was slow. Therefore, the functional protrusion of mandible should be careful.2 The experiment has proved that under the treatment with insulin, the functional protrusion of mandible of diabetic rats can reach effective treatment effects.
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