| Research Background and ObjectiveCurrently, the incidence of diabetes mellitus (DM) has shown a sharp upward trend and seriously affected human health and economic development both in developed and developing countries. Pathogenesis of DM relates to the interaction between heredity and environment, where environmental factors, such as pressures from living and working environment, obesity resulting from unhealthy diet and excessive intake of sugar and fat, bad lifestyle of sedentary habit, can exert an important effect on the onset of DM. The specific mechanism of DM remains unclear, but impaired insulin secretion and insulin resistance are major factors leading to DM. DM-induced acute and chronic complications are the main causes of increased DM harmfulness. With advances in medical technology, the incidence and harmfulness of acute DM complications has decreased significantly, and vascular diseases-based chronic complications, including atherosclerosis, coronary heart disease, cerebrovascular disease, and diabetic nephropathy, diabetic retinopathy and diabetic foot, become the leading causes of the disability and death in DM patients. As a major risk factor for vascular diseases, DM is associated with 2-5 higher incidence of vascular diseases compared with non-DM patients, and 80% of DM patients die from vascular diseases. Atherosclerosis not only acts as an important foundation for diabetic vascular diseases but also appears to be their primary pathological process. Therefore, it is of great significance for the prevention from diabetic vascular hazardous events to identify diabetic vascular diseases as early as possible and carry out timely interventions.Atherosclerosis is an important foundation of diabetic vascular diseases. Many factors may result in atherosclerosis, in which inflammation and abnormal fat metabolism plays a vital role. Hypersensitive C-reactive protein (hsCRP), as the sensitive marker of inflammation and tissue damage, is an acute phase protein produced by liver under the stimulation of IL-6, TNF-a and other inflammatory factors. Numerous studies demonstrated that hsCRP is closely related to atherosclerosis, and pathological observation found CRP deposition in plaques. Through binding to phosphocholine expressed by oxidized LDL, CRP can increase macrophage absorption of LDL, upregulate the expression of endothelial cell adhesion molecules, inhibit the expression of nitric oxide synthase (NOS) of aortic endothelial cells, promote the expression of plasminogen activator inhibitor-1, stimulate vascular endothelial cells to release endothelin-1, IL-6 and monocyte chemoattractant protein-1 (MCP-1), and upregulate NF-kB and induce the expression of VSMCs angiotensin Ⅱ receptor type 1, thus facilitating the increased production of angiotensin Ⅱ-related reactive oxygen species and the occurrence of endothelial cell injury, and finally leading to atherosclerosis. As a result, as the sensitive marker of inflammation factors, hsCRP has a very important role in the formation process of atherosclerosis. Studies have already proven the excellent predictive value of hsCRP for vascular hazardous events. It has also been shown that the hsCRP level in DM patients appears to be higher than the normal population and that hsCRP may also be involved in the pathogenesis of DM. Chronic inflammation can stimulate the body’s stress, induce the massive release of inflammation factors and thus contribute to insulin resistance, which, however, serves as a risk factor both for DM and atherosclerosis. Consequently, monitoring of serum hsCRP levels has a positive significance in predicting the complications of DM and vascular diseases.Heart-type fatty acid binding protein (H-FABP) is the most widely distributed subtype in the fatty acid binding protein family and is expressed in cardiomyocytes, skeletal muscle, aortae, endothelial cells and islet cells; the highest content is reported in myocardium, followed by skeletal muscle. The main function of H-FABP, as the main carrier of fatty acid functioning and metabolism, is to bind to long-chain fatty acids and aid in the absorption and utilization of fatty acid. Meanwhile, it also participates in the release of certain cytokines and may regulate cell proliferation, differentiation and apoptosis as well. Considering the small molecular weight of 15kD, H-FABP can be immediately released from the cells into the blood once the tissue cells it exists are injured. In cases of myocardial injury, H-FABP appears earlier in the blood than the traditional cardiac enzymes and has been now regarded as a marker of myocardial injury. Studies have shown that H-FABP can be a good indicator for the risk stratification of most acute coronary syndromes; H-FABP can be used to predict cardiovascular hazardous events, and the higher the H-FABP level, the higher the probability of cardiovascular events. Few studies available abroad indicate that the serum H-FABP level in DM patients is significantly higher than the normal population, and that H-FABP may be involved in DM by affecting insulin sensitivity and fatty acid metabolism and participate in the formation of atherosclerosis as an inflammation-related factor.H-FABP and hsCRP are recognized predictors of vascular events and are mainly used for the diagnosis and prognosis evaluation of myocardial infarction, cerebral infarction and acute coronary syndrome currently. However, the studies on their association with DM and early vascular diseases have been rarely reported. To this end, by detecting the H-FABP and hsCRP levels and taking carotid intima-media thickness as a reference, their relations with DM and vascular complications have been explored in this paper, hoping to provide valuable detection indexes for the early prediction and intervention of diabetic vascular diseases.Materials and Methods1. Experimental subjectsSubject to the DM diagnosis and typing criterion issued by World Health Organization (WHO) in 1999,88 cases of T2DM patients admitted to our hospital from November 2013 to May 2014 were divided into three group according to the carotid intima-media thickness (IMT) measured by color Doppler ultrasound:1) Carotid IMT-normal group (T2DM-A):28 Type 2 DM patients (15 males,13 females) with IMT<1.0mm and the average age of 51.11±7.22 years; 2) Carotid IMT-thickened group (T2DM-B group):30 Type 2 DM patients (16 males,14 females) with 1.0mm≤IMT<1.5mm and the average age of 51.57±8.47 years; 3) plaque group (T2DM-C group):30 Type 2 DM patients (18 males,12 females) with carotid IMT>.5mm and the average age of 54.53±9.79 years. At the same time,30 healthy subjects (17 males,13 females) that underwent examination in our hospital were selected as the normal control group (NC group) with the average age of 50.03±10.25 years.2. Experimental Methods2.1 Specimen CollectionAfter 12h of fasting, all subjects underwent venous blood drawing before 8:00 of the next morning using two 5ml heparin anticoagulation blood-collection tubes and one 5ml EDTA anticoagulation blood-collection tube. After standing for 30min, they were placed into the centrifuge and centrifuged for 10min at 3000r/min. After that, plasma from either of heparin anticoagulation tubes was transferred to a cryogenic vial and stored at -80℃. for future batch detection of H-FABP.2.2 Experimental MethodsThe serum H-FABP level was detected by ELISA, where the kit was provided by the Lanzhou Institute of Biological Products Co, Ltd. and the specific experimental steps referred to reagent instructions, and each H-FABP concentration was finally calculated based on specimen absorbance. Fasting plasma glucose (FPG), triglyceride (TG), total cholesterol (TC), low-density lipoprotein cholesterol (LDL-C), high density lipoprotein cholesterol (HDL-C), fasting insulin (FINS), hsCRP and glycosylated hemoglobin (HbAlc) were determined by our Clinical Laboratory using automatic analyzers; the insulin resistance index was determined using homeostasis model assessment index HOMA-IR=FBG x FINS/22.5. The carotid intima-media thickness was detected in all research subjects by the doctors of our Ultrasound Department using high-resolution color Doppler ultrasound, where IMT< 1.0mm was defined as normal,1.0mm≤IMT<1.5mm was defined as carotid intimal thickening, and local IMT≥1.5mm was defined as the formation of atheroma.2.3 Statistical MethodsSPSS13.0 statistical software was used for the statistical analysis. The measurement data was represented in the form of (x±s); One Way ANOVA was used to compare the means among groups; the comparison between the two groups was performed using LSD-t test; Dunnett’s T3 was used for pairwise comparison in cases of heterogeneity of variance; count data were analyzed using chi-square test.Results1. Comparison on serum hsCRP and H-FABP levels between each DM group and the normal control groupThe hsCRP levels in NC group, T2DM-A group, T2DM-B group and T2DM-C group were 1.10±0.82 mg/L,1.96±1.20 mg/L,3.62±2.61 mg/L and 5.95±3.76 mg/L respectively with the highest concentration of hsCRP reported in the T2DM-C group, indicating statistical significance compared to other groups (P<0.05). The difference was also statistically significant when T2DM-B group was compared with T2DM-A group (P<0.05), while the difference was not statistically significant when T2DM-A group was compared with the control group. With the increase of IMT value, H-FABP levels, which were 4.15±1.14ng/ml,6.28±3.71ng/ml,8.76±3.74 ng/ml and 11.75±6.67 ng/ml in each group respectively, were also elevated gradually, showing that the differences between each group and the control group were statistically significant (P<0.05) and the differences among three DM groups were also statistically significant (P<0.05).2. Correlation analysis between various biochemical indexes and hsCRP and H-FABPCorrelation analysis results showed that hsCRP was positively correlated with SBP, FPG, LDL-C and HbAlc (P<0.05) and had no correlation with DBP, BMI, FINS, HOMA-IR, TQ TC and HDL-C (P>0.05); HFABP was positively correlated with SBP, DBP, FPG, FINS, HOMA-IR, LDL-C and HbAlc (P<0.05) and had no correlation with BMI, TG, TC and HDL-C (P>0.05).3. Correlation analysis between carotid IMT of each DM group and each indexThe spearson correlation analysis between carotid IMT and each index showed that carotid IMT was closely and positively correlated with course of disease as well as LDL-C, hsCRP and H-FABP levels, suggesting statistically significant differences (P<0.001) with respective correlation coefficients of 0.537,0.348,0.590 and 0.601; carotid IMT was positively correlated with SBP, FINS and HOMA-IR with correlation coefficients of 0.265(P=0.013),0.225(P=0.016) and 0.234(P=0.025) respectively; IMT was not correlated with other indicators. Assessment on regression analysis curve indicated a significant linear correlation of IMT with hsCRP and H-FABP levels (P<0.001).4. Analysis of risk factors for DM carotid IMT using multiple linear regressionIn the DM groups, multiple linear regression analysis was conducted with IMT as the dependent variable and all other indicators as independent variables. The results showed that hsCRP, H-FABP, course of disease and LDL-C eventually came into the regression equation (model) with the model multiple correlation coefficient R=0.778, the coefficient of determination R2=0.606, respective regression coefficients of 0.351,0.274,0.283 and 0.202, and all corresponding P values of less than 0.05, suggesting a remarkable statistical significance.5. Analysis of H-FABP and hsCRP evaluation performance on carotid plaqueThe ROC curves of hsCRP and H-FABP were used to analyze and evaluate their diagnostic capabilities for carotid plaque formation. The results showed that the area under hsCRP receiver operating characteristic (ROC) curve was 0.791 and that at 95% confidence interval was (0.693～0.886, P<0.001), whereas the area under H-FABP ROC was 0.763, and that at 95% confidence interval was (0.671～0.841, P <0.001); the sensitivity of elevated hsCRP on the carotid plaque formation of Type 2 DM was 66.7%, the specificity was 75%, the positive predictive value was 47.6% and the negative predictive value was 86.6%; the sensitivity of elevated H-FABP on the carotid plaque formation of Type 2 DM was 73.3%, the specificity was 68.1%, the positive predictive value was 44.4% and the negative predictive value was 88.2%; the sensitivity of simultaneously elevated H-FABP and H-FABP on the carotid plaque formation of Type 2 DM was 64.5%, the specificity was 83.3%, the positive predictive value was 64.5% and the negative predictive value was 87.3%.Conclusion1. H-FABP and hsCRP are positively correlated with carotid IMT of Type 2 DM patients; elevated H-FABP and hsCRP levels are risk factors for IMT, suggesting that hsCRP and H-FABP may participate in the occurrence and development processes of Type 2 DM atherosclerosis.2. hsCRP and H-FABP are good indicators in predicting Type 2 DM plaques. The combined detection of them has a certain diagnostic value for diabetic vascular plaques. |
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