In the present study, we assessed C-IMT in 490 T2D participants. The arginine stimulation test (AST) method was applied as the established measurement to assess insulin secretion in patients. We for the first time demonstrated that insulin resistance and insulin secretion are independent risk factors of C-IMT in men and women with T2D, respectively.

Elevated C-IMT has been demonstrated to be associated with insulin resistance (evaluated by HOMA-IR), glucose intolerance, and higher fasting glucose levels, especially in non-diabetic individuals and even in obese children [9, 13, 17, 19], indicating that early atherosclerosis in prediabetes may be causally linked to endothelial insulin resistance. In fact, hyperinsulinemia resulted from insulin resistance has been reported to cause endothelial dysfunction and atherosclerosis [20]. More importantly, insulin supplementation showed the ability to improve several vascular functions in animal models and people with insulin resistance [21]. On the other hand, Roussel et al. demonstrated that insulin secretion (evaluated by the early insulin response index which was calculated as the ratio of insulin change over the first 30 min of the OGTT, to plasma glucose at 30 min) is also associated with early carotid atherosclerosis in non-diabetic population independently of other risk factors such as insulin resistance [12]. Similarly, β cell function (estimated by HOMA-β) was still significantly related to an increased risk of poor functional outcomes in non-diabetic ischemic stroke patients after adjusting insulin resistance [18]. Furthermore, C-IMT was addressed to be associated with impaired β cell function in non-diabetic people [11]. Together these hinted that defects in insulin resistance and/or insulin secretion may pathophysiologically be connected with the development of carotid atherosclerosis. Given the above evidence in non-diabetic population, we raised the likelihood that this association may also exist in T2D population. Indeed, our data clearly revealed that C-IMT value is significantly correlated with insulin resistance (evaluated by HOMA-IR) and insulin secretion (evaluated by ACR and AIR) in T2D patients. More interestingly, these associations are manifested in a gender-specific manner, which is in line with the previous findings that C-IMT is associated with insulin sensitivity in men, but with fasting plasma glucose in women, respectively [13]. This discrepancy might reflect a gender-specific mechanism involved in the development of atherosclerosis in T2D patients. However, the explanation behind is not fully understood, and therefore, future studies focus on exploring the causal relationship between impaired glucose metabolism and atherosclerosis in different gender groups are needed.

The progression of C-IMT is determined by a plethora of risk factors, such as age, blood pressure, lipids, smoking, obesity, and CRP [22, 23], though we found the C-IMT value is significantly correlated with insulin resistance and insulin secretion in a gender-specific manner. It is noteworthy that diet, exercise, parental history of premature death from coronary heart disease are also associated with carotid atherosclerotic plaques [24], which may confound the interpretation of our results. Therefore, in the present study, we employed stepwise multivariate linear regression analysis to explore the independent C-IMT-correlated parameters in gender-divided subgroups to adjust the potential confounding factors. After excluding the traditional risk factors (waist circumference, systolic BP, LDL-cholesterol, and current smoking), our multivariate regression model revealed the true independent determinants of C-IMT in T2D population in a gender-specific manner. Taken together, these suggest that the risk factors of carotid plaques as well as the mechanisms underlying may differ in gender in T2D individuals. Further studies are needed to explore the more comprehensive relationships between C-IMT and arginine-stimulated insulin secretion and insulin resistance after excluding all the possible variables.

HOMA-IR and HOMA-β are widely used for evaluating insulin resistance and insulin secretion, respectively [18, 25, 26]. It is noteworthy that these two measurements derived from fasting samples are limited to merely reflecting fasting nondynamic conditions in clinical settings. In addition, oral glucose tolerance test (OGTT) and euglycemic-hyperinsulinemia clamp have also been applied to evaluate insulin sensitivity [11,12,13]. In our study, AST was employed to examine the first phase insulin secretion and the reserved function of β cells. It has been established that β cell dysfunction plays a key role in the pathogenesis of diabetes development that leads to defects in glucose-stimulated insulin secretion. However, β cells still retain the act to stimuli by non-sugar substances such as arginine [27]. Arginine is more potent to trigger secretion in β cells than glucose, and hence, can be used to evaluate the reserved function of islet β cells. In fact, hyperglycemic clamp technique is a more accurate application for evaluating islet β cell secretory function [14]. Nonetheless, the high technique requirement and long operative time limit its utility in clinical settings. In contrast, AST is far less technically demanding and can bring reproducible and complementary measures of β cell function [28]. More importantly, in young T2D individuals, AST is beneficial to reflect β cell reserve regardless of disease duration and treatment [29]. At last, it has also been demonstrated that arginine is preferred to glucagon for the assessment of β cell function [30].

T2D is suggested to be linked with atherosclerotic cardiovascular diseases through multifactorial pathways. For instance, HDL-cholesterol levels or the HDL-based makers have been established to be associated with various disorders, including hypertension [31], hepatosteatosis [32], thyroiditis [33], and in particular, diabetes [34, 35]. Importantly, these conditions are also tightly connected with high burden of inflammation that is well known to play a key role in the development of atherosclerosis in T2D patients [36]. In addition, there is evidence that women generally have higher HDL-cholesterol levels than men [37, 38], which is also confirmed by the baseline data in our study population. These together strengthen our results that atherosclerotic cardiovascular disease is gender-specifically associated with T2D.

Clinically, the gender-specific C-IMT values demonstrated by our findings provide a novel means of risk assessment for T2D patients, which may shed light on the personalized T2D treatment. Moreover, our data also highlight the significance of usage of AST-based insulin secretion in clinical settings to evaluate the relationship between T2D and development of atherosclerosis.

The current study cannot determine the causal relationship between arginine-stimulated insulin secretion and C-IMT due to its cross-sectional and unpaired design. In addition, the relatively small sample size may result in reduced statistical power. A larger population from more clinical study sites is warranted in the future. Notably, patients with diabetes may develop various severe vascular complications, which cannot be merely reflected by C-IMT with ultrasound detection. In addition, C-IMT values may also be affected by many other factors, such as premature deaths in family, diet, and exercise, which may confound the explanation of the results. Therefore, more parameters correlated with C-IMT value and/or atherosclerosis occurrence such as percentage of stenosis and peak systolic velocity (PSV) of carotid that indicating vascular plaque formation are of necessity to be included in the analysis model to establish a more comprehensive explanation.