| Background:Uremia,a common clinical syndrome at the end of chronic kidney disease(CKD),has become one of the major chronic diseases that seriously harm human health.At the same time,the incidence of cardiovascular disease(CVD)in patients with uremia was higher than that of other people,especially coronary artery disease.Coronary artery disease has become an important cause of death in patients with uremia.However,its mechanism is not clear,which may be related to the structural and functional abnormalities of ion channels.Our previous studies have shown that: uremic rats coronary artery lesions with large conductance calcium activated potassium channels(BK channel)express cut and transient receptor potential channel 1 C(TRPC1 channel)expression.They play an important role in maintaining membrane potential of vascular smooth muscle cells(VSMCs)and regulating vascular contraction and diastolic function.Among them,when the expression of TRPC1 channel increases,the intracellular flow of calcium ions will increase,causing vascular contraction,leading to hypertension,atherosclerosis and so on.However,inhibiting the expression of TRPC1 channel or using specific channel blockers can reduce the concentration of calcium ions in cells and mediate vascular relaxation.However,there are no relevant reports on the changes of calcium ion concentration and its mechanism in the cells of coronary smooth muscle of uremia.OBJECTIVE:To study the effect of uremia on calcium ion concentration in rat coronary smooth muscle cells and explore the possible molecular mechanism of calcium ion concentration change in rat coronary smooth muscle cells.METHODS:(1)The healthy SD rats with no significant differences in baseline values were randomly divided into two groups: uremia model group: the model was made by the majority of the 5/6 kidneys.Normal group: the sham operation group,only separated the renal capsule.Both two groups of rats from before and after operation,2,6,10,14,18,22 weeks in orbital blood test of serum urea nitrogen(BUN),creatinine(Scr),to determine whether uremic rats model set up successfully,and at the same time measure the rat tail artery systolic blood pressure(SBP)to observe the normal group and model group rats the dynamic change of blood pressure.(2)Rat coronary smooth muscle cells were isolated from normal group and uremia model by acute enzyme digestion.(3)The calcium ion concentration in normal group and uremia model group was detected by immunofluorescence calcium imaging.(4)Using immunofluorescence calcium imaging normal group and uremia model group rats was detected in coronary artery smooth muscle cells join TRPC1 channel blockers SKF96365 two groups of rats after coronary artery smooth muscle cells in the change of calcium ion concentration.RESULTS :(1)Identification of rat model of uremia:(1)Renal function:compared with normal group,uremia model group rats from 2 weeks after the beginning of serum creatinine(Scr)and the increase of urea nitrogen(BUN),serum creatinine values in 22 weeks up to mu(98.46±3.08)mol/L,blood urea nitrogen up to(44.78 ± 0.21)tendency,L group and normal serum creatinine values for mu(24.28±1.22)mol/L,blood urea nitrogen(7.23±0.31)for tendency for L,conform to the requirements of the building.(2)SBP changes: preoperative no difference between the two groups of rats blood pressure,uremia model group rats(n = 20)SBP rise from 2 weeks after beginning,2 weeks uremia model group rats blood pressure is(130.2 + 4.8)mmHg,and normal group(n = 16)in rats blood pressure is(112.4 + 3.2)mmHg,compared two groups was statistically significant difference(P < 0.05);And longer duration of postoperative,the model group blood pressure is gradually increased,in 6,10,14,18,22 weeks in model group blood pressure value(138.4 + /-4.2 mm Hg,respectively,(150.6-6.6)mmHg,(161.4-6.2)mmHg,(167.2-6.4)mmHg,(179.4-7.3)mmHg,and normal contrast group the difference was statistically significant(P < 0.05).(2)The image captured by MetaFluor software in real time shows:(1)Normal group and uremia model of coronary artery smooth muscle cells within the fluorescence signal intensity ratio R(on behalf of the intracellular calcium ion concentration),respectively: 0.4968 + /-0.0212 and 0.0212 + /-0.0208,differences between the two groups was statistically significant(P < 0.05).(2)The fluorescence signal strength ratio of the cells in the normal group of coronary smooth muscle cellswas 1.7426 + 0.021 and 0.7124 + 0.034,respectively,and the difference between the two groups was statistically significant(P<0.05).(3)The ratio of fluorescence signal strength in the cells of the coronary artery smooth muscle cells of the uremic group was 3.1068 + 0.0224 and 1.1246 + 0.0206,respectively.The differences between the two groups were statistically significant(P<0.05).(4)The difference of the fluorescence signal intensity ratio between the two groups was 1.3642 + 0.215 and1.0302 + 0.027,respectively.The difference between the two groups was statistically significant(P<0.05).CONCLUSION:1.The calcium ion concentration in the rat coronary smooth muscle cells increased after uremia.2.The increase of calcium concentration in rat coronary smooth muscle cells during uremia may be related to the up-regulation of TRPC1 channel expression. |
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