| Objective To determine the optimum experimental conditions of the polymorphism of cyclooxygenase 2(COX-2) gene promoter region-765G/C detected by polymerase chain reaction-restriction fragment length polymorphism(PCR-RFLP). Method In the PCR,6 different kinds of annealing temperature gradient were detected from 58℃-63℃,3 kinds of primers concentration(0.4μm/L,0.6μm/L,0.8μm) were detected experimental results were observed from 2% agarose. In the RFLP,3 kinds of PCR reaction mixture quantity(10μl,13μl,15μl) were detected,2 kinds of incision enzyme quantity(10U,5U) were detected; 2 kinds of incubation time(12h,6h) were detected, experimental results were observed from 3% agarose. Result (1)With annealing temperature increasing, non-specific PCR product amount decreased; however, when annealing temperature exceeding 60℃, PCR amplification result become worse than before. (2)When each of primers concentration was 0.4μmol/L, PCR production level was higher than others and less dimer of primers. (3) It was 15μl PCR production amount that was digested could creat a clear digested electropherogram. (4)It was 5U Bsh1236I that could digest completely total PCR production in single digestion system. (5)It was 6 hours that was demanded in incubated reaction at 37℃condition. Conclution The optimum conditions: annealing temperature was 60℃; primer concentration was 0.4μmol/L; the effective RFLP protocol:the system was 30.5μl including 15μl PCR reaction mixture and 5U enzyme were incubated at 37℃for 6 hours. Objective To determine the possible modulating effect of the cyclooxygenase 2 (COX-2) gene promoter region-765G/C polymorphism on the risk of type 2 diabetes mellitus(T2DM) in Kunming Han people. Method A case-control study was conducted in our study; the polymorphism of the cyclooxygenase 2 (COX-2) gene-765G/C was detected by PCR-RFLP method in 200 unrelated Kunming Han T2DM patients and 100 controls. The genotypes and allele frequencies distributions were compared among cases and controls, at the same time, the related characteristics of cases and controls were compared. Results (1) No significant differences were observed in genotype distributions of the-765G/C polymorphism between cases and controls, also in allele frequencies (χ2=0.915, P＞0.05;χ2=1.842, P＞0.05);(2) In T2DM patients, the FPG level,2hPG level,HbAlc% level in those-765GG genotype carriers were higher than others(FPG:8.85±4.71 vs 7.21±2.65 mmol/L, P＜0.05; 2hPG:12.48±6.0 vs 10.50±4.23 mmol/L, P＜0.05; HbA1C%:8.84±2.76 vs 7.94±2.17, P＜0.05). Conclusion Although COX-2-765G/C polymorphism is not significantly associated with a risk of T2DM in Kunming Han people,-765GG genotype appears to be related to higher plasma glucose level in T2DM patients. Objective To determine the possible modulating effect of the cyclooxygenase 2 (COX-2) gene promoter region-765G/C polymorphism on the risk of diabetic nephropathy(DN) in Kunming Han people. Methods The polymorphism of COX-2 gene promoter region-765G/C was detected by PCR-RFLP method in 152 DN patients, in which including 78 microalbuminuria(DN1 group),74 macroalbuminuria (DN2 group);100 free-DN type 2 diabetes and 100 normal controls. The genotype and allele frequencies distributions were compared among different groups, at the same time, the related characteristics of cases and controls were compared. Results (1) No significant differences were observed in genotype distributions and allele frequencies among the groups, except that-765GG genotype in DN patients was significantly higher than free-DN type 2 diabetes(χ2=4.309, P＜0.05).(2) In all DN patients,the diabetic duration(years) in DN1,DN2 and DNO was 4.83±4.13,7.81±5.77, 4.23±4.00(P＜0.05); the levels of DN2 were higher than DN1 in SBP(166.96±30.58 vs 140.77±27.02),DBP(98.38±15.26 vs 87.96±14.47),FPG(8.24±4.49 vs 7.05±2.82 mmol/L),HbAlc%(9.61±2.50 vs 8.30±2.35),FA(363.90±87.05 vs 330.55±66.05μmol/L), TG(2.83±2.44vs 1.99±1.65 mmol/L),UA(360.68±119.67 vs 315.59±95.44μmol/L), (P＜0.05);the levels of DN2 were higher than DNO in SBP(166.96±30.58 vs 140.43±30.44 mmHg),DBP(98.38±15.26 vs 88.18±14.66 mmHg),FA(363.90±87.05 vs 329.42±83.12μmol/L),TC(4.96±1.15 vs 4.41±0.92mmol/L),TG(2.83±2.44 vs 1.75±1.13 mmol/L),UA(360.68±119.67vs295.13±78.91μmol/L) (P＜0.05); (3) In all DN patients,the levels of those with-765GG genotype were higher than others in FPG(7.89±3.95 vs 5.93±1.67 mmol/L)2hPG(11.89±5.19 vs9.21±4.47 mmol/L), (P＜0.05) (4) Logistic regression analysis indicate that-765GG genotype (OR: 3.25,95%CI:1.336-7.901),TG level (OR:1.319,95%CI:1.012-1.718), diabetic duration (OR:1.027,95%CI:1.006-1.142)were risk factors of DN; SBP(OR:1.038, 95%CI:1.012-1.065) and HbA1C%(OR:1.341,95%CI:1.107-1.625) were risk factors for development of DN. Conclusion The COX-2 gene promoter-765GG genotype,TG, diabetic duration may associated with diabetic nephropathy in Kunming Han people; SBP and HbAlc% may contribute to DN development. Objective To determine the possible modulating effect of the cyclooxygenase2 (COX-2) gene promoter region-765G/C polymorphism on the risk of diabetic retinopathy (DR) in type 2 diabetes mellitus(T2DM) in Kunming Han people. Method The polymorphism of the cyclooxygenase 2 (COX-2) gene-765G/C was detected by PCR-RFLP method in 121(DR) T2DM patients with DR,44(DR0) free-DR T2DM patients and 100(NC) normal controls.The genotypes and allele frequencies distributions were compared among cases and controls, at the same time, the related characteristics of cases and controls were compared. Results (1) No significant differences were observed in genotype distributions of the-765G/C polymorphism among all groups, also in allele frequencies (χ2=1.377, P＞0.05;χ2=0.446,P＞0.05);(2) The diabetic duration (7.66±5.10 vs 5.24±5.93years),HbA1c% (9.52±2.51 vs 7.77±1.92) and TG(2.47±2.33 vs 1.85±1.03mmol/L) in DR were higher than DR0(P＜0.05);(3)Logistic regression analysis indicate that diabetic duration(OR: 1.319,95%CI:1.355-1.669) and HbAlc%(OR:1.427,95%CI:1.173-1.736) were risk factors of DR. Conclusion COX-2 gene promoter region-765G/C polymorphism is not associated with a risk of DR in Kunming Han T2DM patients; in T2DM patients diabetic duration and HbA1C% may contribute to DR. Objective To determine the possible modulating effect of the RANTES gene promoter-28C/G polymorphism on the risk of pre-diabetes in Wa people of Cangyuan county Lincang city Yunnan province. Methods The polymorphism of RANTES gene promoter-28C/G was detected by PCR-RFLP method in 129(PD) Cangyuan Wa pre-diabetes case and 91 (NC) controls. The genotypes and allele frequencies distributions were compared among cases and controls, at the same time, the related characteristics of cases and controls were compared; A Logistic regression analysis was used to estimated markers of pre-diabetes risk; The genotypes and allele frequencies distributions were compared between Wa people and Han people. Results No significant differences were observed in genotype distributions of the-28C/G polymorphism between cases and controls, also in allele frequencies; female (OR:2.927,95%CI:0.170-0.688),the age(OR:1.026,95%CI:1.003-1.051),TG(OR:2.091 95%CI:1.303-3.355),TC(OR:16.338,95%CI:3.880-68.801),LDL-C(OR:20.525, 95%CI:5.610-75.092) were risk factors of PD in Wa people. Conclusion RANTES gene promoter-28C/G polymorphism is not significantly associated with a risk of pre-diabetes in Cangyuan Wan people; Sex, age and levels of TG, TC, LDL-C,HDL-C were associated closely with pre-diabetes. |
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