Alternation Of Intestinal Microbiota And Intestinal CYP3A And P-gp Expression Of Diabetic Rats

ObjectiveThe pharmacokinetic changes in patients with diabetes are generally believed to be caused by the changes of drug metabolic enzymes and transporters.The present studies mostly focus on the change of liver metabolic enzymes and transporters under diabetic condition,the study for intestinal metabolic enzymes and transporters is not enough,relatively.The study aims to investigate the CYP3A and P-gp expression in different intestional segments of rats with type 1 diabetes,along with the alternation of the diversity of bacteria by using Illumia High Throughput Sequencing Technology.The study also aims to determine the effects of diabetes on oral pharmacokinetics of cyclosporine.MethodsRat model of type 1 diabetes was induced by intraperitoneal injection of streptozotocin?STZ,65 mg/kg?.After 5 weeks,the protein expression of CYP3A and P-gp in different intestinal segments?duodenum,jejunum,ileum and colon?of rats were measured by Western blot assay.The mRNA expression of CYP3A and Abcb1was detected by QT-RCR assay.The intestinal contents of jejunum,ileum and colon of rats were collected under aseptic conditions,and the structure of intestinal flora in intestinal segments of rats was analyzed by Illumina high throughput sequencing technology.Rats with type 1 diabetes were induced by intraperitoneal administration of 65 mg/kg streptozotocin?STZ?.Oral pharmacokinetics of cyclosporine?CsA,10mg/kg?was compared between diabetic and normal rats on the 35th day after STZ injection.Concentration of CsA in whole blood was measured by enzyme multiplied immunoassay technique?EMIT?.ResultsThe protein expression of CYP3A and P-gp in the duodenum and jejunum of rats were not affected by diabetes,but the protein expression of CYP3A and P-gp in the ileum and colon of diabetic rats were significantly decreased.The expression levels of CYP3A and P-gp in ileum were 56.5%and 67.8%?P<0.05?in normal rats,respectively.The expression levels of CYP3A and P-gp in colon were 56.5%and71.2%in normal rats?P<0.01?,respectively.Similarly,compared with normal rats,the mRNA expression of CYP3A and Abcb1a/b in the duodenum and jejunum of diabetic rats did not change,but the mRNA expression of CYP3A and Abcb1b in ileum and colon were significantly decreased.The mRNA expression levels of CYP3A1 and Abcb1a in ileum were 10.7%and 50.5%respectively in normal rats?P<0.05?.The expression levels of CYP3A1 and Abcb1a in colon were 1.9%and 48.5%?mRNA?in normal rats,respectively?P<0.05?.Compared with normal rats,the chao1 values of ileum increased significantly in diabetic rats.Other Alpha diversity index?ace value,shannon value and simpson value?had no significant change.In the level of Genus,the Lactobacillus in the ileum of diabetic rats reduced,while the level of Clostridium_sensu_stricto and Turicibacter levels increased.Compared with control rats,the levels of Blautia and Dorea in the colon of diabetic rats decreased significantly,while Turicibacter,Flavonifractor,Marvinbryantia,and Elusimicrobium levels increased significantly.Type1diabetes rats were successfully induced as the fasting blood glucose levels exceeded 11.1 mM after STZ injection for 1 week.Beside glucose,total cholesterol?TC?and triacylglycerol?TG?in serum of diabetic rats were significantly higher than those in normal rats after STZ injection for 35 days.Unlike normal rats,the pharmacokinetics of CsA in diabetic rats exhibited a double peak after oral administration.The T_max and C_max were lower in diabetic rats than normal rats,although without statistical differences(T_max:3.3±1.6 vs 3.2±2.5 h,C_max:579.0±208.5vs 453.0±104.8 ng/mL in normal and diabetic rats).Compared with normal rats,AUC and T_1/2 of CsA in diabetic rats decreased significantly,which were 0.51 and 0.70times those of normal rats,respectively?P<0.01?(AUC:7343.2±2333.7 vs3729.7±1106.6 h*ng/m L,T_1/2:8.5±1.5 vs 6.0±0.9 h in normal and diabetic rats).Conclusions1.Diabetes Mellitus exhibits the synchronous and regional effects on the CYP3A and P-gp expression in the intestinal tract of rats.2.Diabetes Mellitus affects the diversity of bacteria in ileum and colon of rats,possibly due to the alternation on the mechanism of CYP3A and P-gp expression.3.The changes of the oral pharmacokinetics of CsA in rats with type 1 diabetes were not correlated with CYP3A and P-gp in intestine,but the safety of using the drug as clinical medications for diebetic patients should be considered.