Diabetic Angiopathies
Conditions
Brief summary
To explore the possible implications of HLA-DRB1\*04 alleles in patients with type 2 DM and macroangiopathy
Detailed description
Previous studies examining the molecular etiology of diabetes mellitus in China and abroad have mainly focused on the relationship between HLA and type 1 diabetes mellitus (DM) (1-3). Due to the complexity of type 2 DM heredity, few researchers have studied the correlation between type 2 DM and HLA. In recent years, it has been suggested that the development of coronary heart disease (CHD) in diabetic individuals is not merely a complication of diabetic macroangiopathy. CHD and diabetic macroangiopathy share some genetic associations. Previous studies have demonstrated that certain HLA\*DRB1\*04 subtypes are associated with an increased risk of cardiovascular disease. Experimental methods: For genomic DNA extraction, 3-5ml venous blood samples were collected from all patients. Coagulation was prevented using EDTA. The procedure for PCR amplification was as follows: denaturing for 100s at 96°C, annealing for 60s at 63°C, 1 cycle; denaturing for 10s at 96°C, annealing for 60s at 63°C, 9 cycles; denaturing for 10 s at 96°C, annealing for 30 s at 59°C; extending for 30 s at 72°C, 20 cycles; maintaining at 4°C. Analyzing PCR products stained by Ethidium Bromide on 2.5% agarose gel electrophoresis. Bands were observed using an ultraviolet light and a biological imaging system. Statistical method: The degree of agreement with Hardy-Weinberg equilibrium was determined using the χ2 test. A t-test was employed for comparison of CRP levels (M ± s) and a χ2 test was performed to compare allelic frequencies. The relative risk rate (RR) was calculated as (Number of patients with the gene×Number of people in control group without the gene) divided by (Number of people in control group with the gene×Number of the patients without the gene). PC was calculated using Fisher's method if χ2 \> 3.84.
Interventions
OTHERGroup A
150 healthy blood donors without a family history of diabetes mellitus were enrolled in the study.
OTHERGroup B
200 cases of type 2 DM without complications were enrolled in the study.
OTHERGroup C
120 cases of type 2 DM with macroangiopathy complication were enrolled in the study.. Among the group C subjects, 65 individuals had CHD; 55 patients had cerebrovascular disease (CVD); and 30 subjects had a combination of peripheral vascular diseases (PVD) and CHD.
Sponsors
First Affiliated Hospital of Harbin Medical University
Study design
Observational model
CASE_CONTROL
Time perspective
PROSPECTIVE
Eligibility
Sex/Gender
ALL
Age
20 Years to 60 Years
Healthy volunteers
Yes
Inclusion criteria
* clinical CHD (such as angina pectoris, myocardial infarction) diagnosed by dynamic electrocardiogram and ultrasonic cardiogram; * coronary atherosclerosis confirmed by coronary angiographic examination; * cerebral infarction diagnosed by cerebral CT; * common carotid artery intimal-media thickness (IMT) ≥1.2 mm measured by Doppler ultrasonic examination; * extensive irregular stenosis of a lower extremity artery (diameter \< 3mm) or segmentally obstructed.
Exclusion criteria
* age \< 20 years * age \> 60 years
Design outcomes
Primary
| Measure | Time frame | Description |
|---|---|---|
| Test of allelic frequencies | At recruitment | A commercially available kit was used for extraction of genomic DNA from the whole blood samples (PEL-FREEZ Inc., USA). Thirty-two pairs of sequence-specific primers (SSP) for HLA-DRB1\*04 alleles were purchased from One Lambda, Inc. (USA), and the Taq enzyme was purchased from Promega Corp. (USA). The PCR-SSP technique was employed to determine the HLA-DRB1\*04 alleles for each subject. Genes were amplified using the 5700 PCR thermal cycler manufactured by Applied Biosystems Inc. (USA). |
Secondary
| Measure | Time frame | Description |
|---|---|---|
| Evaluation of serum CRP(C-reactive protein) levels | At recruitment | A commercially available 96-well plate ELISA assay kit (introassay CV 1.7%\ 3.9%, interassay CV 2.8%\ 5.1%) was used to quantify hs-CRP in serum (DSL Inc., USA). All samples were evaluated in duplicate. The absorbance (OD) was determined using a BIO-RAD2550 microwell plate reader (USA). |
Outcome results
None listed