| Objective:(1)To explore the relationship between salivary amylase(sAA)activity and susceptibility to type 2 diabetes mellitus(T2DM);(2)To identify the secretion of saliva and sAA in T2 DM rats;(3)To explore the specific mechanism of the obstacle of salivary and sAA secretion in T2 DM rats from pathological lesions and salivary secretion regulatory pathways.Methods: SD rats were randomly divided into normal group(8 rats)and type 2 diabetes mellitus group(T2DM group,22 rats).After T2 DM rats were fed with high-fat and high-sugar diet for 8 weeks,rats in the T2 DM group were intraperitoneally injected with a small dose of streptozotocin(STZ)at 35 mg/kg to build T2 DM rats model.During the experiment,the daily indexes of drinking water,diet,energy intake,body weight and blood glucose were recorded.Saliva samples before and after stimulation with citric acid filter paper were collected to measure sAA activity;After successful modeling,saliva samples from T2 DM rats and normal rats before and after stimulation with citric acid filter paper were collected to determine saliva flow rate,ion concentration,sAA activity and content,and the total protein content.The protein secretion rate(total protein content/time,mg/min)and sAA specific activity(sAA activity/protein content,U/mg)were calculated;After the experiment,the rats were sacrificed by ether anesthesia,and the parotid gland and submandibular gland tissues of were collected.The contents of blood glucose,total cholesterol(TC),triglyceride(TG),propylene glycol(MDA),and the activity of sAA and total superoxide dismutase(T-SOD)were measured;HE staining was used to observe the pathological changes of salivary glandin each group;RT-qPCR was used to detect the mRNA expression of tumor necrosis factor alpha(TNF-α),interleukin 6(IL-6),beta adrenergic receptor(β1-AR).cholinergic receptor(M3R)and aquaporin 5(AQP5);Western Blot was used to detect sAA and β1-AR protein expression in rat saliva and tissues.Results:(1)2 weeks after STZ injection,T2 DM rats showed significant symptoms of more than three symptoms,which showed that the diet,energy intake,drinking water and blood glucose of T2 DM rats were significantly higher than normal rats(p<0.05),urine volume was significantly higher than that of normal rats,and body weight was significantly lower than that of normal rats(p<0.05).In addition,the insulin content and insulin sensitivity index of T2 DM rats were significantly lower than that of normal rats(p<0.05).The blood glucose of T2 DM rats was higher than 11.1mmol/L after 2h of taking glucose solution,and was significantly higher than that of normal rats(p<0.05);(2)The correlation and segmentation analysis between sAA activity and blood glucose and body weight showed that the sAA activity and ΔsAA(the difference in sAA activity before and after acid stimulation)in normal rats were significantly negatively correlated with blood glucose(p<0.05),while the relationship did not exist in the obesity and T2 DM phase.In addition,the lower the sAA activity and ΔsAA of the rats in normal period,the higher the blood glucose when the rats develop into obesity and T2 DM,and the lower the ΔsAA in the normal period,the higher the blood glucose increase in the later stage of T2 DM.the lower the sAA activity in the normal period,the shorter the time required to model the T2 DM model(p<0.05).There was no correlation between sAA activity and ΔsAA and body weight in experimental rats;(3)The salivary index analysis of T2 DM and normal rats showed that the salivary flow rate,protein secretion rate,sAA activity,sAA protein content and sAA specific activity of T2 DM and normal rats increased significantly after acid stimulation(p<0.05);The salivary flow rate,protein secretion rate,sAA activity and ΔsAA,sAAprotein content and sAA specific activity of the T2 DM rats were significantly lower than those of normal rats(p<0.05).In addition,the sAA activity and sAA protein content of the experimental rats showed a positive correlation trend(r=0.549,p=0.064);(4)Pathological lesions of salivary glands in T2 DM rats showed higher glucose,TC and MDA contents and lower sAA and T-SOD activities in the parotid and submandibular gland of T2 DM rats when compared with normal rats(p<0.05);T2DM rat parotid acinar cells showed obvious deformation,atrophy and vacuolization,the secretion of granules at the top of the acinus was reduced,the ducts were dilated,accompanied by inflammatory cell infiltration.The lesions of serous acinar cells also appeared in the submandibular gland,and the expression of TNF-α and IL-6 in both glands were significantly higher than those in normal rats(p<0.05);(5)mRNA expression of β1-AR,M3 R and AQP5 in the parotid of T2 DM rats,and the expression of β1-AR and sAA protein were significantly lower than those in normal rats(p<0.05).In the submandibular gland of T2 DM rats,except for the M3 R,the expression of other genes and proteins was significantly lower than that of normal rats(p<0.05).Conclusion: Salivary sAA activity and ΔsAA have a certain guidance role in the development of blood glucose and the development of T2 DM.Low sAA activity and ΔsAA can lead to higher blood glucose and susceptibility to type 2 diabetes.Under normal conditions,low sAA activity and ΔsAA have higher blood glucose,higher risk of developing type 2 diabetes,and high-fat and high-sugar diet will increase this risk.In addition,T2 DM rats have abnormal salivary and sAA secretion,which may be related to morphological abnormalities and secretory dysfunction of salivary glands,especially parotid glands. |
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