The correlation between proinsulin, true insulin, proinsulin: True insulin ratio, 25(OH) D3, waist circumference and risk of prediabetes in Hainan Han adults

Purpose Diabetes mellitus is a kind of highly prevalent chronic disease in the world. The intervention measures on the risk factors of prediabetes contribute to control and reduce the occurrence of diabetes. This study aimed to investigate the correlation between proinsulin (PI), true insulin (TI), PI/TI, 25(OH) D3, waist circumference (WC), and risk of prediabetes. Methods In this cross-sectional study, 1662 subjects including 615 prediabetes and 1047 non-prediabetes were recruited. Spearman’s correlation analysis was used to explore the association of PI, TI, PI/TI, 25(OH) D3, and waist circumference with prediabetes. Odds ratios (OR) and 95% confidence intervals (CI) were calculated by logistic regression. Receiver-Operator Characteristic (ROC) curve was used to evaluate the risk of prediabetes. Results Our study showed that FPI, 2hPI, FTI, 2hTI, FPI/FTI, and WC could enhance the risk of prediabetes (OR 1.034; OR 1.007; OR 1.005; OR 1.002; OR 3.577, OR 1.053, respectively; all p< 0.001). Stratified analyses indicated that FPI/FTI associated with an increased risk of prediabetes in men (OR 2.080, p = 0.042). FTI have a weak association with prediabetes risk in men and women (OR 0.987, p = 0.001; OR 0.994, p = 0.004, respectively). 2hPI could decrease prediabetes in women (OR 0.995, p = 0.037). Interesting, the sensitivity (86.0%) and AUC (0.942, p< 0.001) of combination (FPI+FTI+2hPI+2hTI+25(OH) D3+WC) were higher than the diagnostic value of these alone diagnoses. The optimal cutoff point of FPI, FTI, 2hPI, 2hTI, 25(OH) D3, and WC for indicating prediabetes were 15.5 mU/l, 66.5 mU/l, 71.5 mU/l, 460.5 mU/l, 35.5 ng/ml, and 80.5 cm, respectively. What’s more, the combination (FPI+FTI+2hPI+2hTI+25(OH) D3+WC) significantly improved the diagnostic value beyond the alone diagnoses of prediabetes in men and women (AUC 0.771; AUC 0.760, respectively). Conclusion The FPI, 2hPI, FTI, 2hTI, FPI/FTI, and WC significantly associated with an increased risk of prediabetes. The combination of FPI, FTI, 2hPI, 2hTI, 25(OH) D3, and WC might be used as diagnostic indicators for prediabetes.


Introduction
Diabetes mellitus (DM) is one of the most important chronic non-infectious diseases in the world, which has a serious impact on people's lives. China holds the largest number of DM patients in the world. According to IDF statistics in 2013, the number of patients with DM aged 20-79 years in China is about 98 million, which is expected to grow to 143 million by 2035 [1]. Type 2diabetes mellitus (T2DM) is one of the major diabetes. Prediabetes also knew as impaired glucose regulation (IGR), is a statue of abnormal glucose metabolism between normal glucose metabolism and diabetes. The epidemiological survey in 2010 showed that the number of patients with diabetes and prediabetes in China increased to 114 million and 493 million respectively, being accountable for 11.6% and 50.1% of adults [2]. The prevalence rate of prediabetes in the same population is usually higher than diabetes. In addition, people with prediabetes risk factors are most likely to develop diabetes than the general population [3]. Therefore, the intervention measures on the risk factors of prediabetes should be strengthened to control and reduce the occurrence of diabetes as far as possible.
Human proinsulin (PI), the precursor of true insulin (TI), which is comprised of TI and C peptide. PI is synthesized and secreted by pancreatic β cells and is mainly metabolized in the kidney. Only a few PI was released into the blood on physiological conditions, while pancreatic β cells can release PI leading to increased PI in blood under pathological conditions. Roder et al. revealed that the increased PI/TI can be used as a reflection of subclinical β cell dysfunction [4]. Another study noticed that PI/TI positively associated with type-2 diabetes [5]. Thus, PI/TI is an available factor to estimate the degree of pancreatic β cell dysfunction. And the enhanced PI/TI is used as an indicator for predicting diabetes risk. 25(OH) D3 (VD3), a kind of steroid hormone, act as a crucial role in the occurrence of diabetes by protecting pancreatic β cells and increasing insulin resistance of myocardial cells [6,7]. A study indicated that the plasma VD3 level was closely associated with the pathogenesis of diabetes mellitus [8]. At present, most studies have found that VD3 negatively related to the risk of diabetes [9][10][11]. In addition, an increased study suggested that 25-hydroxyvitamin D3 can be used as one of the biochemical markers for predicting diabetes [12][13][14].
Waist circumference (WC) is a traditional body surface measurement index for evaluating central obesity which makes a high predictive value for the incidence of diabetes [15,16].
In summary, several previous studies have focused on the correlation between PI, TI, PI/TI, VD3, WC, and glucose metabolism in DM. However, there is not any survey combined with the above five indicators to screen the high-risk population of diabetes. Therefore, this crosssectional study aimed to investigate the correlation between PI, TI, PI/TI, VD3, WC, and prediabetes adults in Hainan Han Population. We further assessed the diagnostic value of PI, TI, PI/TI, VD3, WC, and their combinations in diagnosing prediabetes. Our present study would give any available information for the prevention and management of diabetes.

Study subjects
In the current study, we selected 1662 residents aged � 18 years old who participated in the National Diabetes Prevalence Survey of the Chinese Medical Association in 2017 to carry out a 75-g oral glucose tolerance test (OGTT). We divided the participants into the healthy group and the prediabetes group according to the ADA diagnostic criteria for prediabetes in 2010 [17]. Prediabetes was defined as 100 mg/dl (5.6 mmol/L) � FPG < 126 mg/dl (7.0 mmol/L) or OGTT 2h PG 140 mg/dl (7.8 mmol/L) � 2hPG < 200 mg/dl (11.1 mmol/L) or 5.7% � HbA1C < 6.4% [17]. All subjects must conform to the following exclusion criteria: 1) Patients with diabetes. 2) Patients with malignant tumors, severe liver and kidney diseases, and metabolic osteopathy. This study was approved by the Ethics Committee of Hainan Affiliated Hospital of Hainan Medical University, and all research experiments were performed depending on the standard protocol of Helsinki's Declaration. Each individual was informed of the purpose of the sample collection and received written informed consent from them before the study.

Anthropometry and metabolic variables
The body weight, height, blood pressure, and waist circumference of all subjects were measured on an empty stomach in the morning. Body mass index (BMI) was calculated by the formula of weight / height-squared (m 2 ). Fasting venous blood samples were collected for detecting metabolic parameters. The level of fasting plasma glucose (FPG), fasting true insulin (FTI), high-density lipoprotein cholesterol (HDL-C), low-density lipoprotein cholesterol (LDL-C), total cholesterol (TC), triglyceride (TG) and blood uric acid (BUA) were detected by an automatic biochemical analyzer. The concentration of 25-hydroxyvitamin D3 (25(OH) D3) was checked with a non-radioactive Enzyme Immunoassay kit. The hemoglobin A1c (HbA1c) was determined by high-performance liquid chromatography (HPLC).

Data analysis
All statistical analysis of this study was performed by the SPSS version 17.0 software. Statistical tests were two-tailed and p< 0.05 indicated statistical significance. All variables except for gender were examined for non-normal distribution. Differences in age and clinical characteristics between healthy and prediabetes groups were analyzed by using the Rank sum test. The difference of gender between the cases and controls was analyzed by χ 2 test. The interrelationship among FPI, 2hPI, FTI, 2hTI, 25(OH) D3, WC, and 2hPG was analyzed by spearman's correlation analysis. We further analyzed the correlation of the indicators with prediabetes stratified by gender. We also assessed the association of FPI, 2hPI, FTI, 2hTI, 25(OH) D3, and WC with prediabetes through univariate and multivariate logistic regression analyses with adjusting for age, sex, WC, BMC, FPG, TG, TC, LDL-C, HDL-C, HbA1c, FPI, 2hPI, FTI, 2hTI, and 2hPI/ 2hTI. Odds ratios and 95% confidence intervals were calculated to evaluate the association. The relationship between indicators and the risk of prediabetes in men and women was carried out by using univariate logistic regression analysis after adjustment for age, WC, BMC, FPG, TG, TC, LDL-C, HDL-C, HbA1c, FPI, 2hPI, FTI, 2hTI, and 2hPI/2hTI. Tests for interaction between indicators and gender were performed using likelihood ratio tests. The receiver operating characteristic curve (ROC) was applied to evaluate the diagnostic performance of PI, TI, PI/TI, 25(OH) D3, WC, and the optimal cutoff points also were calculated in diagnosing prediabetes. The optimal cutoff points, sensitivity, and specificity values for diagnosing prediabetes were determined according to the Youden index. The area under the ROC curve (AUC) was used to compare the ability of PI, TI, PI/TI, 25(OH) D3, and WC to diagnose the risk of prediabetes [18].

PLOS ONE
p < 0.001), which the values of sensitivity and specificity were 86.0% and 90.2%, respectively. The sensitivity and AUC were larger than the diagnostic value of these alone diagnoses. When stratified by gender, we observed that the diagnostic value of the combinations (FPI + FTI + 2hPI + 2hTI + 25(OH) D3 + WC) was significantly better than these alone diagnoses in prediabetes both men (AUC = 0.771, p < 0.001) and women (AUC = 0.760, p < 0.001) ( Table 7).
As previous studies have demonstrated that prediabetes was an impaired glucose regulation state which mainly caused by the pancreatic β cell dysfunction with insulin secretion and hepatic insulin resistance (IR) cell damage [19,20]. Besides, people with prediabetes have a high risk of developing diabetes mellitus, especially T2DM [21]. Numerous researches have indicated that PI, TI, 25(OH) D3, and WC were associated with the occurrence of T2DM [5,[22][23][24][25].
In our study, the association between PI, TI, and risk of prediabetes was explored in Hainan Han adults by using spearman's correlation analysis and logistic regression analysis. We found that FPI, FTI, and FPI/FTI ratio positively correlated with prediabetes. FPI, FTI, and FPI/FTI ratio were significantly associated with an increased risk of prediabetes. These results are in agreement with Pradhan's findings in middle-aged women which showed that FTI can be used for a long-term risk predictor, FPI and FPI/FTI have a strong predictive role in the rapid development to T2DM [5]. In general, it seems that FPI and FTI could be a candidate risk factor in the diagnosis of prediabetes. Besides, we also observed that 2hPI/2hTI negatively relates to prediabetes. And the significant relationship was found between 2hPI/2hTI and prediabetes risk. In general, it seems that FPI, FTI, FPI/FTI, and 2hPI/2hTI could be a candidate risk factor in the diagnosis of prediabetes.
We further investigate the correlation between 25(OH) D3 and prediabetes. It found that 25(OH) D3 was positively related to prediabetes, but it cannot use for risk factors. Aleksandra et al. also indicated that 25(OH) D3 was not an association with the risk of prediabetes in US civilians, however, they showed 25(OH) D3 significantly increased risk of T2DM [26]. However, there was a study that revealed that the lower level of 25(OH) D3 has significantly enhanced the risk of T2DM among Japanese adults, especially in the prediabetes population [27]. These diverse results may be due to the regional difference among them.
Several previous studies suggested that WC is a better predictor for hypertension and T2M, however, there is no report on prediabetes [28,29]. Thus, we also detected the correlation between WC and prediabetes. We found that WC significantly associates with increased risk The ROC curve was finally performed to evaluate the diagnostic value of PI, TI, 25(OH) D3, and WC for prediabetes. Interestingly, the sensitivity (86.0%) and AUC (area = 0.942) of combination (FPI + FTI + 2hPI + 2hTI + 25(OH) D3 + WC) were higher than the diagnostic value of these alone diagnoses.
Our present study has some limitations. Firstly, the sample size of participants is relatively small for an epidemiological survey. More sample size could provide further definitive evidence. Secondly, we haven't explored the correlations stratified by age, and so on, which can be investigated in further work. Despite its limitations, the study certainly provided very useful evidence for a diagnostic indicator of prediabetes.