To study the relationship between different sampling site of region of interest (ROI) and effects of contrast-enhanced ultrasound (CEUS) in normal rabbit kidney and to investigate the optimal location of ROI.

CEUS was performed in twenty normal rabbits using GE LOGIQ E9 machine and a 6-9 MHz probe, after i.v. bolus injection of 1.0 ml/kg contrast agent (SonoVue^?, Bracco, Italy). The qualitative analysis of the corresponding compartment was assessed using low mechanical index (MI) true agent detection contrast mode. Corresponding parameters of the time-intensity curve (TIC): arrival time (AT), time to peak (TTP), Peak intensity (PI) and area under the curve (AUC) were measured using time intensity curve analysis software package (GE). Quantification results of the same sampling site of ROI in the left and right kidney were compared and statistically analyzed. Quantification results of different sampling site of ROI in the right kidney were compared and statistically analyzed. Quantification results of the set of ROI in superficial and deep renal cortex which were measured by two different observers were analyzed by Bland-Altman analysis.

CEUS was performed successfully in all of the rabbits. The TICs of different ROIs in one kidney had a similar trend. The TICs of the same ROI in left and right kidneys had no statistical significance (P>0.05). The corresponding parameters of different ROIs in right kidney had significant difference (AT: F=5.61, P<0.05; TTP: F=4.28, P<0.05; PI: F=145.88, P<0.05; AUC: F=216.72, P<0.05). SNK-q test was used for multiple comparisons. Superficial renal cortex vs deep renal cortex, PI had statistical significance (PI: q=4.43, P<0.05); superficial renal cortex vs renal medulla, PI and AUC had statistical significance (PI: q= 12.69, P<0.05; AUC: q=13.39, P<0.05); deep renal cortex vs renal medulla, PI and AUC had statistical significance (PI: q= 8.41, P<0.05; AUC: q=13.13, P<0.05); superficial renal cortex vs renal hilus, PI and AUC had statistical significance (PI: q=-8.14, P<0.05; AUC: q=-11.67, P<0.05); deep renal cortex vs renal hilus, PI and AUC had statistical significance (PI: q=-12.25, P<0.05; AUC: q=-12.78, P<0.05); renal medulla vs renal hilus, AT, TTP, PI and AUC had statistical significance (AT: q=3.91, P<0.05; TTP: q=3.00, P<0.05; PI: q=-19.58, P<0.05; AUC: q=27.12, P<0.05). For the ROI in superficial renal cortex in right kidney, the ICC for PI measured by 2 observers was 0.987 (95%CI: 0.946 to 0.991), indicating a very strong agreement. Bland-Altman pot showed that the mean difference of PI measured by 2 observers was 0.42 (95%CI: -0.89 to 1.73). For the ROI in deep renal cortex in right kidney, the ICC for PI measured by 2 observers was 0.698 (95%CI: 0.238 to 0.881), indicating a common agreement. Bland-Altman pot showed that the mean difference of PI measured by 2 observers was 1.1 ( 95%CI: -3.3 to 5.5).

Different sampling site of ROIs can affect the results of quantitative analysis. It is important to realize the relationship between the sampling site of ROI and contrast-enhancement effects in microcirculatory quantification. The superficial renal cortex appears to be the optimal location of ROI for tissue perfusion studying in healthy rabbit kidney.