| Objective:Retinol bingding protein 4 (RBP4) is a new adipokine found in the circulation, thought to be secreted mainly by adipose tissue and the liver. The main function of RBP4 is to transport the retinol (vitamin A) to target tissues in the blood circulation. More and more studies indicated that RBP4 was involved in the pathogenesis of insulin resistence (IR), type 2 diabetes mellitus (T2DM) and diabetic macroangiopathy. Vitamin D is a secosteroid, which is obtained from exposure to sunlight and through dietary sources including food and supplements, its role in inflammation, immune, insulin secretion and glucolipid metabolism as well as in bone metabolism has been demonstrated, serum 25-hydroxyvitamin D (25(OH)D) is the main circulating form of vitamin D. Lower extremity arterial disease (LEAD) is a part of peripheral arterial diseases and diabetic macroangiopathy, which seriously affects the patient's functional capacity and quality of life, and its pathogenic mechanism is arteriosclerosis. This study is aimed to explore the relationship among RBP4,25(OH)D concerntrations and LEAD in T2DM patients, and to investigate the effects of vitamin D intervention.Methods:107 patients (63 males,44 females) who were recruited from outpatient were in line with the T2DM diagnosis standard delivered by World Health Organization (WHO) in 1999. They were hospitalized from October 2012 to January 2013 in the Department of Endocrinology in Anhui Provincial Hospital.Ankle brachia index (ABI) for all the subjects were examined by Doppler ultrasonography, and ABI<0.9 indicated the presence of LEAD. They were assigned to a group without complications (Group DM1) and another group complicated with LEAD (Group DM2). Then subjects in Group DM2 were further divided into Group DM3 and Group DM4, patients in these two groups were both treated with hypoglycemic drugs and aspirin, and additionally, patients in Group DM3 were given placebo and patients in Group DM4 were orally administered Vitamin D 1000 IU daily.38 cases of healthy people were chosen as a control group (Group NC). Clinical characteristics including sex, age, diabetes duration, history of smoking, height, weight, hip circumference and waist circumference were collected. Body mass index (BMI) and Waist-hip Ratio (WHR) were calculated as weight divided by height squared (kg/m2) and waist circumference divided by hip circumference respectively. Systolic blood pressure (SBP) and diastolic blood pressure (DBP) were measured by a digital automatic blood pressure monitor. Patients' blood samples were kept frozen at -80? until analysis. Fasting plasma glucose (FPG), total cholesterol (TC), triglycerides (TG), low density lipoprotein cholesterol (LDL-c) and high density lipoprotein cholesterol (HDL-c) were detected by automatic biochemical analyzer. HbAlC was conducted by high-pressure liquid chromatography. Fasting insulin (FINS) was tested by radioimmunoassay (RIA). The homeostasis model assessment for insulin resistance (HOMA-IR) was calculated according to the formula (HOMA-IR= FINS (mU/L)× FPG (mmol/L)/22.5). The level of RBP4 was measured by ELISA. Serum 25(OH)D concentrations were detected via enzyme-linked immunoassay (ELISA), it was indicated sufficiency when 25(OH)D?30 ng/ml, insufficiency when 20ng/ml?25(OH)D<30ng/ml and deficiency when 25(OH)D<20 ng/ml. Clinical symptom score was conducted in Group DM2. Pearson's or Spearman's analysis was used to assess the relationship among RBP4,25(OH)D and other markers. Multiariable logistic regression analysis was applied to show independent predictors for the presence of LEAD.12 weeks later, clinic indexes and ABI were reexamined and clinical symptom score was detected to assess the efficacy in Group DM3 and DM4.Results:1. There were no differences of sex between the 3 groups (P>0.05). Raised FPG, FINS, HbA1C, TC, TG, LDL-c, HOMA-IR, SBP, BMI, WHR, RBP4 and decreased HDL-c,25(OH)D in Group DM1 and DM2 were exhibited as compared with Group NC (P<0.05); Age, duration, RBP4, FPG, LDL-c, SBP, DBP, HbA1C, the rate of smoking increased and HDL-c,25(OH)D, ABI decreased in Group DM2 compared with Group DM1 (P<0.05). The percentages of sufficient 25(OH)D levels in Group DM1 and DM2 were both lower than the percentage in Group NC(P<0.05). Compared with Group DM3, RBP4, HOMA-IR, TG, LDL-c, FINS, clinical symptom score decreased and 25(OH)D, ABI raised significantly (P<0.05) in Group DM4 after 12 weeks of vitamin D supplementation. The total effective rates in Group DM3 and DM4 were 67.74% and 93.55% respectively, the difference was statistically significant (P<0.05).2.Analysis of Pearson correlation revealed that RBP4 was positively associated with TG (r=0.112,P<0.05), LDL-c (r=0.39,P<0.05), HOMA-IR (r=0.436,P<0.05), BMI (r=0.269,P<0.05), WHR (r=0.551,P<0.05) and negatively associated with 25(OH)D (r=-0.538, P<0.05) and HDL-c (r=-0.278,P<0.05).25(OH)D had a negative correlation with RBP4 (r=-0.538, P<0.05), duration (r=-0.663,P<0.05), HbAlc (r=-0.482,P<0.05), HOMA-IR (r=-0.26,P<0.05) and FPG (r=-0.229,P<0.05). Logistic regression analysis showed that FPG, LDL-c, SBP and smoking were risk factors of LEAD while HDL-c, 25(OH)D were protective ones.Conclusion:1. Increased serum RBP4 levels and reduced 25(OH)D concentrations are significantly associated with LEAD occurrence in patients with T2DM.2. The supplementation of vitamin D can reduce the levels of RBP4 and protect T2DM patients against LEAD by improving IR, lipid metabolism and inhibiting inflammation.Objective:Recent investigations showed that low-grade chronic inflammation and insulin resistance (IR) were the common pathological basis of both diabetes and artherosclerosis. Adipocytokines and inflammatory factors are related with the occurrence and development of Type 2 Diabetes (T2DM) complicated with macroangiopathy. As a new kind of adipocytes, Retinol-binding protein 4 (RBP4) are associated with IR and T2DM. Nuclear factor-kappa B (NF-?B) is a kind of transcription factor which participates in inflammation mechanisms of artherosclerosis and regulates gene expressions. Vitamin D is a secosteroid, which is formed in the skin upon UV-light exposure as vitamin D3 or is ingested by diet as vitamin D2, it is associated with inflammation, immune, insulin secretion and glucolipid metabolism along with its classic role in calcium homeostasis and bone metabolism. We aimed to observe the relationship among RBP4, NF-?B and diabetic atherosclerosis and to study the intervention effect of vitamin D by establishing streptozotocin-induced diabetic rats and diabetic atherosclerosis rats models.Methods:75 male SPF Wista rats were randomly divided into four groups:a normal control group (group NC), a group of diabetic rats (group DM1), a group of diabetic rats with atherosclerosis (group DM2) and a group of diabetic atherosclerosis rats treated with vitamin D (group DM3). At the 16th weekend, all the rats were killed and blood samples were gathered from left ventricles of their hearts, some of the samples were used for assay of fasting plasma glucose(FPG), low density lipoprotein cholesterol (LDL-c), high density lipoprotein cholesterol (HDL-c) and triglycerides (TG) by an automatic biochemical analyze, some were used for fasting insulin (FINS) detected by radioimmunoassay (RIA), and the rest were centrifuged to detect serum RBP4 by enzyme-linked immunosorbent assay (ELISA). Atherogenic index of plasma (AIP) and Homeostasis model assessment of insulin resistance (HOMA-IR) were calculated according to the formulas AIP=logTG/HDL-c and HOMA-IR= FINSxFPG/22.5 respectively. The thoracic aortas were separated and removed for HE staining and the measurement of NF-?B activity by ELISA. Adipose tissue in mesenterium, epididymis and peritoneal were taken out for the measurement of the percentage of fat mass, besides, the epididymis adipose tissue were additionally for RBP4mRNA detection by reverse transcription polymerase chain reaction (RT-PCR) analysis. Arteria caudilis systolic blood pressure (SBP) were assayed by a RBP-1 determinator.Results:(1) Compared with group NC, the expression of RBP4 in serum and adipose tissue, the activity of NF-?B in aorta significantly increased in group DM1 and DM2, and the mentioned indexes above further increased in group DM2. Compared with group DM2, above-mentioned indexes significantly decreased in group DM3 after 12 weeks of vitamin D supplementation. (2) Compared with group NC and DM3, TG, LDL-c, FPG, FINS, the percentage of fat mass, SBP, HOMA-IR, AIP significantly increased in group DM1 and DM2 while HDL-c decreased. Compared with group DM1, TG, LDL-c, FPG, FINS, the percentage of fat mass, SBP, HOMA-IR, AIP significantly increased in group DM2. (3) The structure of thoracic aortas in group NC and group DM3 had no significant pathological changes. In group DM1, the surface of aortic tunica intima was not smooth and glossy, the arrangement of vascular endothelial cells was irregular, the structure of medial smooth muscle was disorder, few foam cells could be seen diffusively. In group DM2, the aortic tunica intima was thickened and protruded into the lumen, the arrangement of the smooth muscle cells was irregular, the muscle fibers were rupture, amount of foam cells, atherosclerotic plaques and calcifications could be seen concentratedly. (4) Pearson Correlation analysis showed that serum RBP4 was positively associated with TG (r=0.329, P<0.01), LDL-c (r=0.399, P<0.05), FINS (r=0.562, P<0.05), HOMA-IR (r=0.663, P<0.01), the percentage of fat mass (r=0.550, P<0.01), AIP (r=0.432, P<0.01), NF-?B (r=0.481, P<0.01), SBP (r=0.449, P<0.01) and negatively associated with HDL-c (r=-0.363, P<0.01); RBP4mRNA expression in adipose tissue was positively associated with TG (r=0.498, P<0.01), LDL-c (r=0.362, P<0.01), FINS (r=0.326, P<0.01), percentage of fat mass (r=0.470, P<0.01), weight (r=0.317, P<0.01), AIP (r=0.429, P<0.01), HOMA-IR (r=0.371, P<0.01), NF-?B (r=0.371, P<0.01), SBP (r=0.561, P<0.01) and negatively associated with HDL-c (r=-0.421, P<0.01), there is a positive correlation between serum RBP4 levels and RBP4mRNA expression in adipose tissue (r=0.633, P<0.01). The presence of atherosclerosis was used as dependent variable, and the rest of factors were used as independent variables in T2DM groups, multivariable logistic regression analysis showed that serum RBP4 (OR=1.409,95%CI=1.188-2.305) and TG (OR=1.179,95%CI=1.093-1.475) were significant predictors.Conclusion:(1) The expression of RBP4 in serum and adipose tissue, the activity of NF-?B in aorta were increased significantly in group DM1, and the metioned indexes above were further increased in group DM2. (2) Increased RBP4 and NF-?B levels were significantly associated with the occurrence of diabetic atherosclerosis. RBP4 might involve in the occurrence of atherosclerosis by IR, inflammation and glucolipids metabolism disorders. (3) The supplementation of vitamin D can reduce the expression of RBP4 and the activity of NF-?B, it had protective effects against diabetic atherosclerosis, which is partially associated with the relief of excessive inflammation, the improvement of lipid metabolism and the elevation of insulin sensitivity. |
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