The Effect Of Uridine On Blood Glucose Level And Liver Metabolism And Its Mechanism Exploration

BackgroundAbnormal glucose metabolism is a common metabolic disorder in obesity,diabetes,pancreatic and liver diseases.Hyperglycemia are the main cause of the deterioration of these diseases and a series of complications.Long-term hyperglycemia often causes irreversible damage to other vital organs such as the heart,brain,kidney,eyes and so on.Currently,a large number of drugs to control blood glucose have been applied in clinical treatment,but most of them have more or less side effects.It is of great significance to explore new compounds that can regulate blood glucose with less adverse effects and the mechanism may provide a new direction for the treatment of diseases related to abnormal glucose metabolism.Uridine is a small molecule nucleoside which widely exists in blood,cerebrospinal fluid and other body fluids and all kinds of tissues.Studies have shown that uridine has important physiological effects on reproductive system,circulatory system and nervous system,and has been applied to reduce the side effects caused by cancer chemotherapy.In recent years,researches indicated that uridine could improve glucose tolerance in high-fat diet mice and aging mice,and this effect is closely related to the effect of leptin.However,the effect and mechanism of uridine on normal blood glucose mice and other types of abnormal glucose metabolism mice have not been studied.The liver is one of the most important organs for regulating glucose homeostasis,and the effect of uridine on liver metabolism has not yet been fully investigated.And the level of uridine in human plasma and its correlation with blood glucose levels have not been systematically observed and studied.This study will focus on these three aspects to explore the role of uridine in these processes.Part Ⅰ: The effects of uridine on blood glucose level and its possible mechanismObjective: To study the effects and mechanisms of uridine on blood glucose level in mice.Methods: To construct an acute treatment model for intraperitoneal injection of uridine in C56BL/6 mice.The effects of uridine on blood glucose in fasting and nonfasting mice were observed and the effects of different uridine doses on blood glucose changes in mice were also investigated.The effects of uridine on blood glucose in mice with different duration of high-fat diet(HFD)and the effect of uridine on glucose tolerance of HFD mice were studied.Changes in plasma insulin levels after uridine treatment were detected.Observing the effects of uridine on blood glucose in streptozotocin(STZ)-induced type 1 diabetes model mice and leptin deficiency(ob/ob)mice.To observe the effect of N-(phosphonacetyl)-L-aspartate(PALA)on blood glucose in mice.Realtime-q PCR was used to detect the effects of insulin synthesis and secretion-related gene expression in mouse pancreas treated with uridine.To observe the effects of uridine and P2Y6 receptor inhibitor(MRS2578)on insulin secretion of mouse islet cell line MIN6 in vitro.Results: The blood glucose level of non-fasted male C57 mice was significantly reduced(blood glucose decreased by 2.00 ± 0.95 mmol /l,P<0.05)after 2h of intraperitoneal injection of uridine,and the blood glucose level of female C57 mice was also significantly reduced(blood glucose decreased by 1.68 ± 0.45 mmol /l,P<0.05).Moreover,uridine could cause a decrease in blood glucose level in fasting mice(6.65 ± 0.76 mmol/l,0h → 4.75 ± 0.95 mmol/l,2h,P<0.001).The blood glucose level of mice decreased significantly 0.5h after the intraperitoneal injection of uridine,and decreased to the lowest level about 1h after the injection,followed by a slow rise in glucose level,and then returned to the pre-injection level at 3h.The subjects of 0.25mg/g uridine administration could cause a significant decrease in blood glucose level(blood glucose decreased by 0.93 ± 0.52 mmol/l,P<0.05).Uridine also reduced blood glucose levels in HFD model mice at HFD-1 day,HFD-1 week,HFD-2 weeks,and HFD-16 weeks.Uridine also improved glucose tolerance in HFD – 16 week mice[AUC values in PBS group and uridine group were 2243 ± 98.69 mmol/(l·min)and 1768 ± 70.64 mmol/(l·min),respectively].However,uridine could not change blood glucose levels in STZ-induced type 1 diabetic model mice and ob/ob mice.Uridine causes a brief rise in plasma insulin levels in mice and an increase in the expression of P2Y6,a gene associated with islet secretion.In vitro culture of MIN6 cells,the application of P2Y6 receptor inhibitor can reduce the increase of insulin level stimulated by uridine.Conclusions: Uridine could rapidly reduce blood glucose levels in normal mice and high-fat diet mice,and the mechanism of this hypoglycemic effect may be related to the increased insulin secretion promoted by uridine.Part Ⅱ: Study on the effect of uridine on liver metabolism and its possible mechanismsObjective: To explore the effect and mechanism of uridine on liver metabolism.Methods: C57BL6 mice were divided into the uridine treatment group and the control group and were intraperitoneally injected with uridine and PBS,respectively.Liver tissues and plasma of the mice were collected 1 hour after uridine administration.The liver tissues were used for periodic acid shiff(PAS)and Oil red staining.Liver glycogen,liver glucose,hepatic triglyceride(TG)and total cholesterol(Tch)were measured.To detect the expression of glucose metabolism related genes in liver.Plasma glucagon levels in mice were also measured.The whole transcriptome sequencing was performed on the liver using the Next-generation sequencing technology.Results: PAS staining showed that the liver glycogen of uridine group was significantly less than that of control group.The glycogen content in the uridine group was also significantly lower than that in the control group(29.61 ± 10.49 mg/g vs 56.35 ± 11.90 mg/g,P=0.0038).There was no significant difference between the two groups in liver Oil red staining and the contents of hepatic TG and Tch.The plasma glucagon level of the uridine group was significantly higher than that of the control group(191.28 ± 15.73 pg/ml vs 108.18 ± 17.37 pg/ml,P<0.001).The liver glucose content of uridine group was also significantly higher than that of PBS group(30.58 ± 4.80 μmol/g vs 18.00 ± 1.93 μmol/g,P<0.001).Transcriptome data analysis showed that uridine had significant effects on transcription factor activity,extracellular membrane formation,ribonucleoprotein,circadian rhythm and epithelial cell differentiation.It also has obvious effects on carbohydrate phosphatase activity,hexose metabolism and lipid metabolism.The expression of genes related to glucose metabolism in liver showed that the expression of genes related to gluconeogenesis and glycolysis,G6 PC,G6PC2,Gck and Pfkfb3 were significantly increased,while the expression of Pfkfb1 was significantly decreased.Conclusions: Uridine could promote the decomposition of glycogen in the liver in a short time,and this process may be related to the increase of glucagon level.Uridine also causes the liver to accumulate large amounts of glucose,which may be one of the mechanisms by which uridine reduces blood sugar and may be related to the increased expression of Gck in the liver.In addition to the changes in glucose metabolism,uridine may also affect the activities of transcription factors,extracellular membrane formation and altered the ribonucleoprotein,circadian rhythm and epithelial cell differentiation.Part Ⅲ: The correlation between fasting serum uridine level and obesity and blood glucose level in humanObjective: To explore the factors affecting serum uridine level in human samples and the correlation between serum uridine and obesity and blood glucose.Methods: Routine health data were from 503 men and women(age>18)participating in an annual physical checkup at Renmin Hospital of Wuhan University(Wuhan,China)in 2019.Fasting serum uridine(FSU)and glucose concentrations and other blood parameters were determined after an overnight(> 12h)fast.Height,weight,and waist circumference were objectively measured.Covariates included age and sex while stratification analysis.Results: In cross-sectional analyses,the mean FSU level was 7.018 ± 1.429 μM for the entire population(N=503).The obese(BMI ≥ 28 kg/m2)group showed significantly higher FSU than the normal weight group(17.5 ≤ BMI < 20)(7.411 ± 1.487 versus 6.662 ± 1.323 μM,P = 0.0107).FSU was positively correlated with BMI(r = 0.183;P < 0.0001)and waist circumference(r = 0.163;P < 0.0001),but negatively correlated with fasting glucose(r =-0.184,P < 0.0001).Uric acid,the purine nucleotide degradation product that is often viewed as a counterpart of uridine,showed a positive correlation with BMI(r = 0.357,P < 0.0001),but no correlation with fasting glucose,highlighting that the inverse correlation of FSU and glucose is specific and BMI-independent.Conclusions: FSU is associated with obesity in individuals with well-controlled fasting glucose;however,a high level of fasting uridine is associated with a hypoglycemic effect when the BMI is comparable,suggesting uridine as a BMIindependent determinator of fasting glucose.