IVIM DWI Evaluation In The Experimental Study Of CIAKI And Initial Clinical Application Of Diabetic Nephropathy

[Object]To investigate the feasibility of IVIM DWI (intravoxel incoherent motion diffusion weighted image) in evaluating the dynamic change of renal functions both in normal and CIAKI rats’ kidney. And to further compare the efficiency among IVIM DWI for diagnosing CIAKI.To further study the clinical value of IVIM DWI in evaluating the renal injury with DN (Diabetic Nephropathy)[Materials and Methods]1. Animals and equipmentsTwenty-seven male Sprague-Dawley (SD) rats with body weights of 200-250g were obtained from Sun yat-sen university animal experiment center. All animals were kept under standard conditions and fed with standard rodent chow and free water. All the rats were anaesthetized with an intraperitoneal injection of 20% urethane (6 ml/kg b.w.). The CIAKI rat model were obtained by administrating ionic iodinated contrast agent (Meglumine Diatrizoate,370mg/ml,6 ml/kg) via tail vein injection.6 CIAKI rats were selected to perform MR exam, the CIAKI rats were scanned 24 h before and 30 min,12 h,24 h,48 h,72 h and 96h after the injection of ionic iodinated contrast agent. Each rat was fasted for solids and liquidsfor 4 hours before MRI scans. The group before CM injection was regarded as the control group. At each of the 7 MR examination time points,3 rats were executed to get right kidney specimens for HE staining.All experiments were performed using a 3.0-T whole-body system (Signa EXCITE HD, GE Healthcare, Milwaukee, WI) with a 40mT/m (200 mT/m-ms) gradient system and a transmit birdcage body coil with 50-mm outer diameter and an eight channel receive-only volumetric rat array (RAPID Biomedical GmbH, Rimpar, Germany), for homogeneous RF transmission and signal detection respectively. The rats were placed head first and supine to make the kidneys in the RF center of the rat array coil. We use abdomen tamping to fix the rats. To determine the dynamic change of intra-renal perfusion and oxygenation.Ethics:All procedures were approved by the Guangdong Academy of Medical Sciences Research Ethics Committee.2. experimental patients and healthy volunteersDiabetic nephropathy group enrolled 53 patients with diabetic nephropathy were from guangdong general hospital.4 cases were excluded because of poor image quality that affect the data measurement.3 cases were excluded for severe atrophy of kidney. According to the eGFR (estimated glomerular filtrationrate), patients were divided into three groups:the early DN group included 18 cases (eGFR> 60 ml·min-1· (1.73 m2)-1), with trace or large amount of proteinuria,10 men,8 women, age ranged from 43-73, the median age was 60 years; the intermediate stage DN group included 15 cases (eGFR> 30 ml·min-1·(1.73 m2)-1,<60ml·min-1· (1.73 m2)-1), with medium or large amount of proteinuria,10 men,5 women, age ranged from 31-74, the median age was 60 years old; terminal stage DN group included 13 cases (eGFR<30 ml·min-1·(1.73 m2)-1), with a large amount of proteinuria,10 men and 3 women, age ranged from 44-69, the median age was 55.Control group enrolled 17 healthy volunteers,11 men,6 women, age ranged from 44-76, the median age was 55. inclusion criteria:people with no history of chronic diseases such as hypertension, diabetes, blood urea nitrogen and serum creatinine are normal.All participants signed informed consent.3. MRI examination (1) MR scanning series for ratMRI scanning sequences including conventional axial T1WI, axial and coronary T2WI sequence, and I VIM DWI functional imaging sequences. The details of the pulse sequences used are listed below:(a) FSE-XL/T1WI sequence:Axial, Scan Mode=2D,Grad Mode=zoom, TR=500 ms, TE=13.3 ms, Flip Angle=90°, Bandwidth=15.6 Hz, NEX=4.0, FOV=10.0×7.0 cm, Slice Thickness=2.0 mm, Sap=0.2 mm, Matrix=288×192, ET=3, Scan Time=94 s.(b) FSE-XL/T2WI sequence:Axial, Scan Mode=2D, Grad Mode=zoom, TR=4500 ms, TE=125.8 ms, Bandwidth=62.50 Hz, NEX=6.0, FOV=10×7.5cm, Slice Thickness=2.0 mm, Sap=0.2 mm, Matrix=288×192, ET=19, Scan Time=443 s.(c) FSE-XL/T2WI sequence:coronal, Scan Mode=2D, Grad Mode=zoom, TR=4500 ms, TE=125.8 ms, Bandwidth=62.50 Hz, NEX=6.0, FOV=10×7.5cm, Slice Thickness=2.0 mm, Sap=0.2 mm, Matrix=288×192, ET=19, Scan Time=443 s.(d) In IVIM DWI, ten b values (0,20,40,60,80,100,200,400,500 and 600 s/mm2) were applied with a single-shot diffusion-weighted spin-echo echo-planar sequence. A local shim box covering the kidney region was applied to minimize susceptibility artefacts. In total, three coronal slices covering the kidney were obtained with a 10-cm field of view,2.4-mm slice thickness,0.2-mm slice gap,3,000-ms TR, 73.6-ms TE,96^96 matrix and NEX=8. Scan Time=4 min 06s.(2) MR scanning series for humanMRI scanning sequences including conventional axial T1WI, axial and coronary T2WI sequence, and IVIM DWI functional imaging sequences. The details of the pulse sequences used are listed below:(a) FSE-XL/T1 WI sequence:Axial, Scan Mode=2D,Grad Mode=zoom, TR=6.18 ms, TE=2.84 ms, Flip Angle=20°, Bandwidth=41.67 Hz, NEX=2.0, FOV=40.0cm x40.0 cm, Slice Thickness=8.0 mm, Sap=2.0 mm, Matrix=288×192, ET=1, Scan Time=1 min 41 s.(b) FSE-XL/T2WI sequence:Axial, Scan Mode=2D, Grad Mode=zoom, TR=6000 ms, TE=102 ms, Bandwidth=62.50 Hz, NEX=2.0, FOV=38cm x 38cm, Slice Thickness=5.0 mm, Sap=2.0 mm, Matrix=288×224, ET=13, Scan Time=2 min 54 s.(c) FSE-XL/T2WI sequence:coronal, Scan Mode=2D, Grad Mode=zoom, TR=15789.5ms, TE=240 ms, Bandwidth=50 Hz, NEX=2.0, FOV=42cm×42cm, Slice Thickness=3.0 mm, Sap=0.2 mm, Matrix=320×192, ET=29, Scan Time= 3 min 10 s.(d) In IVIM DWI, ten b values (0,20,40,60,80,100,200,400,500 and 600 s/mm2) were applied with a single-shot diffusion-weighted spin-echo echo-planar sequence. A local shim box covering the kidney region was applied to minimize susceptibility artefacts. In total, three coronal slices covering the kidney were obtained with a 32-cm field of view,3.0-mm slice thickness,0.6-mm slice gap,3,000-ms TR, 60.3-ms TE,96×96 matrix and NEX=8. Scan Time= 2 min 06s.4. Post-processingAll MR images were evaluated by two radiologists with over 10 years experience blindly to the group designations. The two radiologists were asked to reach a consensus if there was a discrepancy in the result. All images were used to measure ADC ADC, D, D*and f values by functool-MADC software on Advantage Workstation 4.3 GE Medical System (AW4.305). We selected right kidney to position regions of interest (ROIs). Under the condition of anesthesia and special coil, we got high image quality of CIAKI rats, and the kidney can be divided into cortex (CO), outer medulla (OM) and inner medulla (IM). Because of the influence of breathing and heart beating, the kidney of Patients and healthy volunteers can be divided into cortex and medulla, ADC, D, D*and f values were obtained including cortex (CO), outer medulla (OM) and inner medulla (IM). The ROIs were selected at the superior pole, inferior pole and the level of renal hilum of the kidney, with the same size and shape. For rat, each ROI had an area of 2.5-5.0 mm2 and contained at least five pixels. ROI was measured in cortex, outer medulla, and inner medulla. For human, each ROI had an area of 30-50 mm2 and contained at least five-ty pixels. ROI was measured in cortex and medulla and without any distortions, artifacts, large vessels, renal collecting system or any incidental renal cysts in the ROIs.5. pathological examinationAt each time point, we executed 3 rats after MR scanning to get right kidney specimen for pathological examination, The kidney specimens were preserved in 10% formalin for fixed preparation, section, stain and observed under 100 and 200 times of the optical microscope. According to the positive area expression, we divided the area into three sections:strong positive area, positive area and weakly positive area. Then we extracted the positive region from images by using video camera, and measured grey value of the region.6. Statistical analysisAll datas were analyzed using SPSS 19.0 for Windows. One-way ANOVA test were used for comparing ADC, D, D* and f values of cortex, outermedulla and innermedulla for rat among each time. LSD test was used for comparison within class. Nonparametric test was used when the distribution was non-normal or the variance was heterogeneous. P<0.05 was used as the criteria for the statistical difference between two groups.The results of DN patients and healthy volunteers in the form were recorded as mean±standard deviation. One-way ANOVA test were used for comparing ADC, D, D* and f values of cortex and medulla among each group control group, the early stage DN group, intermedian stage DN group and terminal stage DN group. LSD test was used for comparison within class. Nonparametric test was used when the distribution was non-normal or the variance was heterogeneous. P<0.05 was used as the criteria for the statistical difference between two groups.[Results]1. Measurement and comparison of IVIM DWI parameters for CIAKI Rats at each time pointThe time-signal curve slope of cortex and was slightly greater than outermedulla ewspecially in low b value range (0-200 s/mm2).In CO, a progressive reduction was observed in D and ADC by 3.07%(P=0.014) and 8.62%(P=0.001) at 30 min, and the minimum D and ADC value occurred at 48 h. A later changed pattern was found in OM and IM, where D and ADC values were significantly smaller (12-72 h) than baseline, followed by a progressive increase starting from 96 h. A significant decrease was shown for fin both the cortex and medullar in an early stage of CI-AKI (12-48 h) and then ascend in the later stage (72-96 h). D* was decreased by 26.02%(P= 0.143),21.78%(P=0.143) and 10.19% (P= 0.143) in CO, OM and IM at 30 min. The recovery for D* was found at 48 h, which was also prior to f in CO and OM.2. Measurement and comparison of IVIM DWI parameters for DN patients and healthy volunteersTo compare and analysis the D and ADC values that associated with diffusion, the result shows that in the early stage DN group, the ADC values in cortex and medulla(2.26±0.04×10-3 mm2/s,2.02±0.03×10-3 mm2/s) appear significant lower (P<0.05) than control group(2.71±0.03×10-3 mm2/s,2.34±0.02×10-3 mm2/s), and the D values in cortex and medulla(1.71±.06×10-3 mm2/s,1.48±0.06×10-3 mm2/s) appear significant lower (P<0.05) compared with control group(2.24±0.08×10-3 mm2/s,2.71±0.03×10-3 mm2/s). In intermediate stage DN group, the ADC values in cortex and medulla (1.70±0.03×10-3 mm2/s,1.52±0.02×10-3 mm2/s) and D values in cortex and medulla (1.49±0.06×10-3 mm2/s,1.25±0.04×10-3 mm2/s) show significantly lower (P<0.05) than early stage DN group. In terminal stage DN group, the ADC values in cortex and medulla (1.36±0.04±10-3 mm2/s,1.22±0.04×10-3 mm2/s) and the D values in cortex and medulla (1.04±0.04×10-3 mm2/s,1.07±0.03× 10-3 mm2/s) appear significant lower (P<0.05) compared with intermediate stage DN group; D values in three groups always show significant lower than ADC values (P< 0.05), ADC values and the D values in cortex were significant higher than in the medulla (P<0.05).To compare and analysis the D* and f values that associated with perfusion, the result shows that in the early stage DN group, the D* values in cortex and medulla(30.1±14.95×10-3 mm2/s,38.0±11.87×10-3 mm2/s) appear significant lower (P<0.05) than control group(43.1±10.80×10-3 mm2/s,45.4±10.7×10-3 mm2/s), and the f values in cortex and medulla(31.6±19.59%,30.1±14.3%) appear significant higher (P<0.05) compared with control group (26.1±10.03%,26.9±7.82%). In intermediate stage DN group, the D* values in cortex and medulla (23.9±7.83×10-3 mm2/s,28.0±17.24×10-3 mm2/s) and f values in cortex and medulla (21.4±16.74%, 23.6±13.22%) show significantly lower (P<0.05) than early stage DN group. In terminal stage DN group, the D* values in cortex and medulla (18.6±9.12×10-3 mm2/s,20.9±13.59×10-3 mm2/s) and the f values in cortex and medulla (14.8±8.88%, 18.8±7.64%) appear significant lower (P<0.05) compared with intermediate stage DN group; in the three groups, D* values in cortex always show significant lower than in the medulla (P<0.05).[Conclusions]According to the study of CIAKI rats, IVIM DWI can effectively appear the dynamic change process of kidney function after CI-AKI, partlv reveal the micromechanism of CI-AKI. D value reflect the water diffusion state inkidney, D* value depends on expanding or contracting state of the renal capillary, while the f value reflects the liquid load of kidney to some extent. The first part of this study has made a beneficial exploration for clinical application with IVIM DWI in CI-AKI even non cancerous lesions of kidney. According to the part of DN patients research, IVIM DWI can effectively display dynamic change process of kidney function with diabetic nephropathy, that reflects the micromechanism of pathological changes of diabetic nephropathy. The second part of the study had shown that IVIM DWI can make noninvasive assessment of kidney function with diabetic nephropathy, providing reference for clinical diagnosis and therapy, consequently, IVIM DWI appears potential value of clinical application.