Ethical statement
The Human Subjects Committee of the Affiliated Changzhou No.2 People′s Hospital with Nanjing Medical University approved this study. Written informed consent was obtained from each patient enrolled to the research. The reference number for the ethical approval is: [2014]KY010–01.
Study population
Fifthy-seven untreated T2DM patients (37, males) and 52 normal controls (27, males) of similar age and gender were enrolled for this study, however, 4 untreated T2DM patients (2, males) were excluded for the large difference in heart rate, 1 untreated T2DM patients (male) was excluded for fat, and 1 untreated T2DM patients (female) was excluded for COPD. At last, 51 untreated T2DM patients (34, males) were enrolled for the study (Fig. 1). This is a cross-sectional study that was conducted between August 2015 and May 2017, and data was collected prospectively. Subjects with cardiovascular disease, such as coronary artery disease (all of the patients were had coronary artery CT scan to ensure that they had no coronary artery disease), arterial hypertension, myocardial infarction, cardiomyopathy, valvular disease, atrial fibrillation, congenital heart disease, thyroid disease, neoplastic disease, or kidney failure were excluded from the study. In normal control subjects, all of the physical examination tests, the electrocardiogram, and the echocardiography were showed normal. All normal subjects had an absence of diabetes (according to the diagnosis of T2DM, the fasting blood glucose, Two-hour postprandial blood sugar and HbA1c were all showed normal).
Biochemistry
In T2DM patients and normal subjects, fasting plasma glucose, two-hour postprandial blood sugar and glycated hemoglobin (HBA1c) were measured.
Conventional 2D Doppler echocardiography
All enrolled subjects underwent conventional 2D Doppler echocardiography (Vivid E9, GE healthcare). Echocardiography examination was done before beginning drugs in T2DM patients. Left atrial diameter (LAD), interventricular septum thickness and LV posterior wall thickness in end-diastole (IVSD and LVPWD) were measured in the parasternal long-axis view of LV by M-mode. LV end-diastole volume (LVEDV), LV end-systole volume (LVESV) and LV ejection fraction (LVEF) were measured by modified biplane Simpson′s method. Peak early and late diastolic velocities of mitral valve (E and A, respectively) were measured by pulsed-wave Doppler, and the ratio of E/A was then calculated. Peak early (e′) and late (a′) diastolic annular velocities were obtained by averaging the values at septum and lateral positions using pulse wave Tissue Doppler Imaging (TDI), and E/e′ was calculated.
ECG leads were connected to each patient. Standard high frame rate (> 60 /s) of the apical four-, three- and two-chamber views of three consecutive cycles while patients held their breath were stored for offline analysis.
Analysis of LV systolic function
The apical four-, three- and two-chamber views were analyzed using 2D-STE software (2D-Strain, EchoPAC PC 113, GE Healthcare, Horten, Norway) by two experienced cardiologist.
First, we defined cardiac PSLR as the rotation of the LV cross section. Segmental and global PSLR degrees were measured. Using the SAX-MV option of EchoPAC software displayed on the apical four-chamber view. The LV walls were divided into six segments: base-septal, middle-septal, apex, middle-lateral, and base-lateral, and one segment containing mitral valve, respectively. The segment containing mitral valve was excluded for the analysis. Then segmental and global PSLR of LV walls were measured via the software (Fig. 2).
Second, we used LAX, A4C and A2C options for the analysis of LV longitudinal strain and strain rate by the software, “LAX” means apical three chamber view. LV longitudinal strain (including LS-endo, LS-mid, and LS-epi, which represented LV endomycardial, middle myocardial and epimycardial walls, respectively) and LV longitudinal strain rate (LSr) were calculated and recorded.
Statistical analysis
All data analyses were performed using SPSS 21.0 software (SPSS, Chicago, IL, USA). Data was presented as the mean ± standard deviation (SD). p-value < 0.05 was considered statistically significant in all tests. Kolmogorov-Smirnov′s test was used to detect the normality of all the segmental and global PSLR values. Differences between T2DM patients and normal subjects were compared with an independent Student′s t-test for the data distribution was normal. Differences among the first analysis, interobserver and intraobserver were compared with one-way analysis of variance (ANOVA). For variables with a non-normal distribution, the nonparametric U Mann-Whitney test was used. Chi square was used for comparing the variable of sex. Spearman′s correlation was chosen for the test correlations among the fasting plasma glucose, HBA1c, LVEF, LS-endo, LS-mid, LS-epi, LSr, segmental PSLR and global PSLR. We defined the apex and global PSLR in control subjects as the normal state, and considered the values of T2DM patients as abnormal. Values for apex and global PSLR in T2DM patients were determined from receiver operating characteristic (ROC) curve analysis. Yoden′s index was selected for the cut-off point which can give the best composite of specificity and sensitivity.
Reproducibility and repeatability
Intraobserver and interobserver variability for global PSLR were determined by repeating measurements in all enrolled T2DM patients and normal subjects. For the second intraobserver measurements, the observer was “blinded” to results of the initial measurements.