Effects Of Inflammatory Cytokines IL-1β And IFN-γ On Insulin Secretion In Rats With Pancreatic Islet Glucose Stimulation

Objective:To investigate the isolation and purification of pancreatic islets in rat SD rats.Methods:The pancreatic islets were isolated and purified by Histopaque 1077 density centrifugation and manual extraction.The islet production and purity were calculated after dithizone(DTZ)staining.The insulin activity was evaluated by insulin secretion response(GSIS).Results:(1)The average purity was 85%and the survival rate of islet cells was 90%in each SD rats.(2)The insulin secretion was(5.77)in the blank control group(10.14 ±1.67)μIU/ml,(14.52 ± 0.75)μIU/ml,(10.58 ± 0.60),respectively,under the stimulation of 5.5,11,1,16.8 and 22.2 mmol/L glucose concentration,)MIU/ml and(7.87.52 ±0.65)μIU/ml,and the blank control group had statistically significant difference(F = 30.56,p<0.05)compared with each group.Conclusion:The pancreatic islets were digested by retinal perfusion of collagenase NB8 in the bile duct,and the islets were isolated and purified by Histopaque1077 density centrifugation and manual extraction.The purity and purity of the waslets were higher.Isolated rat pancreatic islets were treated with different concentrations of glucose,and the insulin secretion was the highest at 11.1 mmol/L glucose stimulation.Objective To investigate the effects of inflammatory cytokines IL-1β and IFN-γ on insulin secretion in rats with pancreatic islet glucose stimulation.Methods Rat waslets were isolated and purified.After 24 hours of culture,the rat waslets were plated with 12-well plates(20 holes each),and the wells were well-rounded.IL-1β(0.25 ng/ml,2.5 ng/ml,25 ng/ml)and/or IFN-γ(10 u/ml,100 u/ml,1000 u/ml)for 24 h,0.5%BSA in KRBH buffer for 1 hour and incubated for 1 hour in 0.5%BSA-containing KRBH buffer containing 11.1 mmol/L glucose.Islet supernatant was collected and the level of insulin secretion was measured by radioimmunoassay.Results:The insulin concentration in the blank control group was(11.25 ±3.73)uIU/ml,and the insulin secretion of the glucose-stimulated group was(40.39 ± 4.12)ulU/ml,IL-1β(36.04 ± 3.35)uIU/ml and IL-1β(2.5ng/ml)group were treated with 11.1 mmol/L glucose concentration at the concentration of 11.1 mmol/L glucose stimulation(29.27 ± 2.33)uIU/ml,IL-1β(25ng/ml)group was(26.84 ± 2.11)uIU/ml with 11.1mmol/L glucose stimulation,and 11.1 mmol/L glucose stimulation group(P<0.05).Thedifference was statistically significant(F = 58.92,p<0.05).IL-1β(25ng/ml)group had the most significant inhibition of GSIS in rat pancreatic islets,and the insulin release was 33.5%lower than that of 11.1mmol/L glucose stimulation group(26.84 ± 2.11)pairs(40.39 ± 4.12)uIU/ml,(29.27 ± 2.33)vs(40.39± 4.12)uIU/ml,(t = 5.339,p<0.05),and the level of insulin secretion in IL-1β(2.5ng/ml)group was 27.5%lower than that in glucose-stimulated group(36.04 ± 3.35)pairs(40.39 ± 4.12)uIU/ml,(t = 2.303,p>0.05).The level of insulin secretion in IL-1β(0.25ng/ml)group was 10.7%lower than that in glucose-stimulated group In the IFN-y(10u/ml)group,the insulin secretion was(42.99 ± 3.99)uIU/ml and the IFN-y(100u/ml)group at 11.1mmol/(30.63± 1.14)uIU/ml,IFN-γ(1000u/ml)group was(23.31± 3.05)uIU/ml,11.1mmol/L glucose concentration at 11.1mmol/L glucose-stimulated glucose There was significant difference between the stimulation group and the IFN-γ group(F = 104.1,p<0.05).IFN-γ(1000u/ml)group had the most obvious inhibition of GSIS in rat pancreatic islets,and the secretion of insulin was 48.4%(23.31± 3.05)(45.19± 3.26)uIU/ml,(30.63± 1.14)pairs(45.19 ± 3.26)uIU/ml,(t=1 1.99,p<0.05),and the level of insulin secretion in IFN-γ(100u/ml)group was 32.2%lower than that in glucose-stimulated group 0.05).The secretion of insulin in IFN-y(0.25ng/ml)group was 10.7%lower than that in glucose-stimulated group(42.99± 3.99)(45.19± 3.26)uIU/ml,(t= 2.43,p<0.05).The levels of insulin secretion were(16.22± 2.44)uIU/ml with 11.1 mmol/L glucose stimulated by IL-1β and IFN-γ,and there was significant difference between the two groups(F = 86.04,p<0.05).(16.22 ± 2.44)vs(26.84 ± 2.11)uIU/ml,(t = 7.554,P<0.05),and the inhibitory effect of GSIS was significantly higher than that of IL-1β(25ng/ml)(16.32 ± 2.44)pairs(22.39 ± 2.74)uIU/ml,(t =4.109,p<0.05),and the insulin secretion was decreased by 27.5%compared with IFN-y(1000u/ml)There was significant difference(F = 86.04,p<0.05)Conclusion Inflammatory cytokines IL-1β and IFN-y can inhibit the insulin secretion of pancreatic islet glucose in rats,and they have synergistic effect.Objective:To investigate the effect of inflammatory cytokines IL-1 β and IFN-γ on the expression of islet insulin gene in rat pancreatic islet glucoseMethods Rat pancreatic islets were isolated and purified.After 24 hours of culture,the rat waslets were plated with 12-well plates(60 holes each),and the wells were well-rounded.(2.5 ng/ml,25 ng/ml)and/or IFN-[gamma](10 u/ml,100 u/ml),the rats were washed twice with PBS buffer twice,0.5%BSA KRBH buffer 1ml/well,cultured for 5min;Total RNA was extracted from the pancreas after culture,and the expression of insulin gene was detected by RT-PCR.Results The expression of insulin gene was significantly increased at the concentration of 22.2mmol/L for 5min,and the expression of insulin gene was increased with IL-1 β and/or IFN-γ in rats after hyperglycemia And the inhibitory effect of insulin gene expression was more obvious under the combined action.