| Because of some factors such as economy developing, lifestyle changing and population ageing, the morbidity of diabetes has risen greatly. The data from World Health Organization indicated that there were 30 million patients in 1985, 135 million in 1997, and will be doubled in 2025. Diabetes has become the third lethal disease after the cardiovascular disease and cancer. Acute and chronic complications are the leading cause of death and disability, especially cardiovascular diseases. Research indicated diabetic patients have high risk of atherosclerosis. Coronary heart disease, cerebrovascu 1 ar disease and peripheral vascular disease are more frequent in diatetes than nondiabetes.Atherosclerosis is a complicate process, many cells and factors taking part in it. The mechanism ofatherosclerosis remains to be understood completely. Such as the disfunction of endothelial cell, the aggregation of monocytes and T lymphocytes from peripheral to intima, The transformation of macrophage from monocyte, the migration and proliferation of vascular smooth muscle and so on, which are the important events during the atherosclerosis. The macrophages not only internalize the lipids, but act as a source of growth-regulatory molecules and cytokines as well. In 1991, Higashiyama discovered heparin-binding epidermal growth factor-like growth factor (HB-EGF) secreted by Macrophage-like U937 cell, which belongs to epidermal growth factor family. Study implied that HB-EGF secreted by macrophage played an important role in atherosclerosis by acting as a potent mitogen and chemokine for vascular smooth muscle cells. Extracellular high glucose could enhance the mitogen of HB-EGF to vascular smooth muscle cell. So HB-EGF may be involved in the occurrence and process of atherosclerosis in diabetes.Up to now, there have had few reports about the relationship between the HB-EGF secreted by macrophage and diabetes, or the regulation of macrophage HB-EGF expression in diabetes. In present study, we estimated the mRNA level of macrophage HB-EGF in type 2 diabetes byRT-PCR to research the possible role of HB-EGF in diabetic atherosclerosis.The study group included 44 patients with type 2 diabetes and 34 normal controls, matched with patients for sex, age, and body mass index. Peripheral venous blood was collected by Sodium Citrate anticoagulating tube, then mononuclear cells were obtained by Ficoll. Based on the anchoring characteristic of monocytes to plastic, cells were isolated and cultured within plastic petri dishs containing RPMI 1640.then were differentiated into macrophage supplemented with 20% autologus serum. After 4 days, macrophages were collected and confirmed by immunohistochemistry. Total RNA from cultured macrophages was extracted with Trizol reagent and reverse-transcripted into complementary D.\A by Oligo dT primer. A 605bp HB-EGF product was amplified using a sense 5'- TGG TGC TGA AGC TCT TTC TGG-3' and an antisense 5' -GTG GGA ATT AGT CAT GCC CAA-3' primer pair. A 370bp β-actin acting as internal standard was amplidied using a sense 5' -TCG ACA ACG GCT CCG GCA-3' and an antisense 5' -CGT ACA TGG CTG GGG TGT-3' primer pair. 5μl of each reaction mixture was e 1 ec t r ophor esed on a 2. 0% agarose gel and the bands were confirmed by directly sequencing after purifying. Relative mRNA levels were assessed by KodakDigital Science 1D.The results showed that the macrophage mRNA levels were significantly higher in patients with type 2 diabetes than those in controls (0.876+0.422 vs. 0.473+0.210 p<0.05). Furthermore, the macrophage mRNA levels were positively correlated with fasting blood glucose (r=0.342 P=0.002).These data suggested that macrophage HB-EGF mRNA was over expression in patients with diabetes, which was related to the hyperglycemia in diabetic state. |
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