Relationship between omentin-1 and carotid intima thickness in type 2 diabetes mellitus

Introduction Omentin-1 is a novel adipokine that has a pivotal role in modulating insulin sensitivity, immunity, and inflammation. Adipokines contribute directly to the atherosclerotic process. The current study was conducted to evaluate the serum omentin-1 level in type 2 diabetes mellitus (DM) patients and to evaluate its relationship with carotid intima media thickness (CIMT). Patients and methods Sixty participants were enrolled in the study: 30 patients with type 2 DM and 30 controls with normal glucose levels. Patients were classified into group I, which included 15 patients with CIMT greater than 0.9 mm, and group II, including 15 patients with CIMT less than 0.9 mm. All groups were subjected to full medical history taking, clinical examination, and assessment of fasting blood glucose levels, lipid profile, and serum omentin-1 levels using enzyme-linked immunosorbent assay. BMI and CIMT were also assessed using color Doppler ultrasound. Results Serum omentin-1 levels were significantly decreased in patients with type 2 DM compared with controls and were further decreased in patients with increased CIMT. Omentin-1 levels were negatively correlated with fasting blood sugar, BMI, waist circumference, lipid profile, and CIMT, and were positively correlated with high-density lipoprotein, with r-values of −0.72, −0.9, and −0.81 for fetal bovine serum, BMI, and CIMT, respectively; −0.58, −0.70, and −0.49 for triglyceride, low-density lipoprotein, and cholesterol, respectively; and +0.66 for high-density lipoprotein. Conclusion Serum omentin-1 level is decreased in type 2 DM patients and is negatively correlated with CIMT and BMI. Hence, omentin-1 could serve as a protective marker and predictor for cardiovascular disease. Further study needed to show whether omentin-1 is considered as a risk factor for DM.


Introduction
Diabetes mellitus (DM) is a widespread health problem. Th e prevalence of DM is expected to rapidly increase from 171 million (2.8% of the world's population) in 2000 to 366 million (4.4% of the world's population) by 2030 [1]. Individuals with DM have two-to threefold increased risk for myocardial infarction or stroke compared with individuals without DM [2].
However, the underlying mechanisms linking type 2 DM with cardiovascular disease remain unclear.
Recently, evidence has shown that some adipokines are major regulators of insulin resistance and direct mediators of endothelial dysfunction and macrophage infi ltration of vessel walls [3].
Mature omentin is secretory glycoprotein consisting of 295 amino acids and n-linked oligosaccharides, and its basic structural unit is a 120 kDa homotrimer in which 40 kDa polypeptides are bridged by disulfi de bonds [4].
Omentin-1 is a new type of Ca +2 -dependant lectin, with an affi nity for galactofuranosyl residue-containing constituents of pathogens and dominant immunogens [5].
containing omentin-1 is added to a well of a microtitre plate preloaded with human omentin-1 monoclonal antibodies and is incubated. Th en add omentin-1 antibodi es labeled with biotin and combined with streptavid in HRP to form Immune Complex.

Measurement of carotid intima media thickness
CIMT is measured manually using a high-resolution B-mode tomographic ultrasound system (MyLab50, Esaote, Italy) with a 7.5-10 MHz linear transducer. Precision of the CIMT measurement is 0.01 mm. CIMT was measured on the fall wall of the right and left common carotid arteries, 1.5 cm proximal to the bifurcation. Th e transducer was manipulated so that the lumen diameter w as maximized in the longitudinal plane. Th e mean CIMT value of the right and left common carotid arteries was used for analysis. A carotid plaque was defi ned as a focal structure protruding into the arterial lumen, with a thickness 1.3 mm or higher; the mean thickness is 0.08 mm.

Body mass index
BMI was calculated as weight/height 2 (kg/m 2 ), and waist circumference (WC) was measured at the midpoint between the lower border of the rib cage and the iliac crest.

Statistical design
Data collected were reviewed. Coding and statistical analysis of collected data were carried out usi ng SP SS program (version 15; SPSS Inc., Chicago, Illinois, USA).
(1) Descriptive statistics: (a) Central tendency and dispersion of quantitative data were expressed as mean and SD. (b) Qualitative data were expressed in terms of frequency of occurrence. (2) Analytical statistics: (a) Groups were compared using the following tests: (i) χ 2 -test: for comparison of qualitative data. (ii) Student's t-test: for comparison of quantitative data between two groups. (iii) Pearson's test: Pearson's correlation coeffi cient was used to determine the association between two variables. (b) Th e level of signifi cance was taken at a P-value of less than 0.05. (c) Th e results are represented in tables and graphs.

Results
Th is study was conducted on 30 patients with type 2 DM and 30 healthy participants. Patients included Th erefore, in this study we measured serum omentin levels in patients with DM to determine whether there is relationship between omentin levels and CIMT, to determine whether serum omentin levels are correlated with other risk factor; for example, BMI, waist circumference, and lipid profi le, and to determine whether omentin-1 can be considered as a protective and predictive marker for cardiovascular diseases in diabetic patients.

Patients and methods
Th e present study is a hospital-based case-control study carried out over a period of 6 months, from December 2012 to May 2013, on a total sample of 60 participants (30 patients with type 2 DM and 30 age-matched and sex-matched controls with normal glucose tolerance levels). Patients were selected randomly from those admitted to the Internal Medicine Department of Al Zahraa University Hospital.
Patients were classifi ed into two groups: Group I: included 15 patients with type 2 DM with CIMT greater than 0.9 mm.
Group II: included 15 patients with type 2 DM with CIMT less than 0.9 mm.
Th irty participants with normal glucose levels served as controls.
Th e patients were subjected to biochemical tests. Fasting blood glucose levels were determined: diabetes was defi ned by a fasting plasma glucose value of 7 mmol/l or higher (126 mg/dl). All participants in the control group were subjected to a 75 g oral glucose loading test to exclude those with prediabetes and those with undiagnosed diabetes. Serum cholesterol, highdensity lipoprotein (HDL), low-density lipoprotein (LDL), and triglyceride levels were determined colorimetrically on Hitachi 736 (Hitachi, Tokyo Japan). Serum concentration of omentin-l was determined usi ng enzyme-linked immunosorbent assay kits.

Principle of the assay
Th e kit assays the level of human omentin-1 in samples on the basis of the double-antibody sandwich enzymelinked immunosorbent assay technique. Th e sample 13 men and 17 women; their ages ranged f rom 39 to 48 years, with a mean ± SD age of 43.20 ± 3.26 years. Th ey were classifi ed as follows: Group I: included 15 diabetic patients (seven male and eight female) with CIMT greater than 0.9 mm; their ages ranged from 39 to 48 years, with a mean ± SD age of 43.73 ± 3.15 years.
Group II: included 15 diabetic patients (six male and nine female) with CIMT less than 0.9 mm; their ages ranged from 38 to 48 years, with a mean ± SD age of 42.67 ± 3.39 years.
Controls: included 12 men and 18 women; their ages ranged from 35 to 47 years, with a mean ± SD age of 41.40 ± 3.31 years.
On comparing the patient group with the control group, an insignifi cant diff erence in age and sex was found between them. However, a signifi cant increase in the fetal bovine serum (FBS) level was found in the patient group (151.87 ± 19.83) compared with the control group (93.73 ± 7.78). In addition, anthropometric measures such as BMI and waist circumference were also signifi cantly higher among patients (30.10 ± 4.44 and 98.20 ± 11.65, respectively) compared with controls (23.47 ± 2.35 and 82.60 ± 5.05, respectively). With regard to the lipid profi le, a signifi cant increase in cholesterol, triglyceride, and LDL levels was recorded in the patient group (179.48 ± 43.07, 173.52 ± 57.82, and 96.55 ± 22.69, respectively) compared with the control group (118.87 ± 33.39, 97.00 ± 12.02, and 32.87 ± 7.31, respectively), whereas the HDL level was signifi cantly lower in the patient group (34.13 ± 6.642) compared with the control group (44.93 ± 6.59). Serum omentin-1 level was signifi cantly decreased in the patient group (6.75 ± 3.30) compared with the control group (11.92 ± 1.34). CIMT was signifi cantly higher in the patient group (0.93 ± 0.20) compared with the control group (0.64 ± 0.08) (  (Table 2).
On comparing patients of group I with controls, there was an insignifi cant diff erence in age and sex between both groups. Th ere was a signifi cant increase in the FBS level in group I patients (160.67 ± 21.48) compared with the control group (93.73 ± 7.78). In addition, BMI and waist circumference were also signifi cantly higher  Table 4).  Omentin-1 levels had a signifi cantly negative correlation with age, fasting blood sugar, BMI, waist circumference, cholesterol, triglyceride, and LDL levels, and CIMT. In contrast, they had a signifi cantly positive correlation with HDL level. Th ese correlations were mild (with age, cholesterol), moderate (with triglyceride and LDL levels, duration of the disease, and BMI), and marked (with FBS, CIMT, and waist circumference; Table 5 and Figs 1-3).

Discussion
Adipose tissue produces several hormones and cytokines termed adipokines, which have widespread eff ects on carbohydrate and lipid metabolism. Th ey appear to play an important role in the pathogenesis of insulin resistance, diabetes, and atherosclerosis [10].
Correlation between serum omentin -I and FBS levels. FBS, fetal bovine serum.

Figure 1 Figure 2
Correlation between serum omentin-1 level and CIMT. CIMT, carotid intima media thickness. Omentin-1 is a newly identifi ed adipokine that is highly and selectively expressed in visceral adipose tissue [5]. It enhances insulin sensitivity and glucose metabolism [4].
Th is study showed a signifi cant decrease in serum omentin-1 levels among diabetic patients in comparison with the normal group (P < 0.00; Table 1), which agreed with several studies that reported that omentin-1 levels in type 2 DM patients are decreased compared with those in healthy controls [11,12].
Th is demonstrates the role of omentin in regulating metabolic function by stimulating glucose uptake in response to insulin in cultured adipocytes, suggesting that omentin has a benefi cial eff ect on insulin sensitivity [4]. In addition, it has been reported that plasma omentin-1 levels declined after prolonged insulin-glucose infusion in healthy individuals. Further, treating the human omental tissue plants with glucose and insulin caused downregulation of omentin-1 expression [11].
We further demonstrated a signifi cant decrease in the omentin-1 level in patients with an increase in CIMT, compared with the other diabetic group. Th is agrees with the fi ndings of Yamawaki et al. [13], who reported that the plasma concentration of omentin is associated with endothelium-dependent vasodilation. In addition, Shibata et al. [14] found that circulating omentin levels were negatively correlated with CIMT, which is a marker of early atherosclerosis. Low levels of omentin are also associated with an increased prevalence of coronary artery disease [15].
Greulich et al. [16]found that omentin-1 was highly expressed and secreted by epicardial adipose tissue, had reduced expression among type 2 DM patients compared with controls, and was positively correlated with diastolic function. Th ese data suggest that omentin may not only serve as a biomarker for metabolic disorders but also as a cardioprotective adipokine, and that a decrease in its level could contribute to the induction of cardiovascular dysfunction in DM patients.
Fantuzzi and Mazzone [3] reported that some adipokines, such as resistin, peptin, and adiponectin, directly mediate vascular health by infl uencing the function of endothelial cells, arterial smooth muscle cells, and macrophages in the vessel wall.
Two other reports on the negative correlation of serum omentin-1 with CIMT have suggested cardioprotective and antiatherosclerotic roles of omentin-1 [17,14]. Hye Jin Yoo et al. [18] suggested that omentin-1 is regulated by infl ammation, which is the most important factor linking type 2 DM with the progression of cardiovascular complications.
An experimental study by Somimaruyama et al. [19] to demonstrate the role of omentin in stimulating endothelial cell function and revascularization processes shows that systemic delivery of an adenoviral vector expressing omentin enhances b lood fl ow recovery and capillary density in ischemic limbs of mice, which is accompanied by increased phosphorylation of Akt (also called protein kinase B) and endothelial nitric oxide synthase. In addition, treatment with omentin protein stimulated the phosphorylation of Akt and endothelial nitric oxide synthase in human umbilical vein endothelial cells, a s well as stimulated the phosphorylation of AMP-activated protein kinase.
We found a signifi cant diff erence in BMI and waist circumference between the diabetic group and the normal group. In addition, there was signifi cant increase in BMI and waist circumference in group I compared with group II.
Th e negative correlation of omentin-1 level with BMI and waist circumference is explained by Desouza Batista et al. [7], who reported that among various human tissues, visceral adipose tissue produces a large amount of omentin and its gene expression in the visceral fat depot is reduced in obese individuals. Pan et al. [12] reported that low levels of circulating omentin are associated with obesity-induced metabolic dysfunction such as insulin resistance and glucose intolerance; this suggests that reduced levels of omentin may be an indicator of visceral fat accumulation, thereby being correlated with clustering of metabolic disorders.
Further, we found a signifi cantly negative correlation between omentin-1 levels and cholesterol, triglyceride, and LDL levels, which agrees with the fi ndings of Reishibata et al. [20], who reported that omentin-1 levels were signifi cantly lower in patients with one

Figure 3
Correlation between ser um omentin-1 level and waist circumference.
or more risk factors of metabolic disorders such as hyperlipidemia.

Conclusion
Th is study showed a signifi cantly decreased omentin-1 level in type 2 DM patients as compared with controls, with a larger decrease among patients with increased CIMT. Th ere was a negative correlation between omentin-1 level and other risk factors for cardiovascular and metabolic disorders such as BMI and lipid profi le. Hence, we can consider omentin-1 levels to be predictive of atherosclero sis and early vascular complications of DM.