Contrast-enhanced ultrasonography is a method emerging in veterinary medicine for the evaluation of tissue and parenchyma perfusion. Contrast agents used in CEUS are composed of gas-filled microbubbles, which are injected into the bloodstream and serve as intravascular reflectors of ultrasound waves to provide real-time assessment of vascular perfusion.1 These contrast media are safe products and are tolerated well by dogs and cats.2
It has been reported that CEUS is accurate for the quantification of liver and splenic perfusion in healthy dogs3–5 and for differentiation between malignant and benign hepatic, renal, and splenic nodules in dogs and cats on the basis of perfusion patterns.6–9 In particular, a rapid influx of contrast agent followed by rapid clearance (early wash-in and early washout) is typically seen in malignant focal liver lesions.10,11 The reliability for differentiation of splenic lesions is lower11–13; however, the assessment of vascular tortuosity may be helpful in discriminating between malignant and benign focal splenic lesions in dogs.14
Contrast-enhanced ultrasonography has been used to distinguish an accessory spleen from a mass of another origin,15 for perfusion imaging of the adrenal glands in healthy dogs and dogs with pituitary-dependent hyperadrenocorticism,16,17 and to investigate the status of prostate gland vascularity in healthy dogs.18 Quantitative CEUS analysis of perfusion in the kidneys, liver, pancreas, spleen, small intestine, and mesenteric lymph nodes of healthy anesthetized cats and dogs has been described.19–24
Several studies25–29 on CEUS analysis of renal perfusion in humans and dogs have been reported. The analysis of data obtained with this technique can require the use of special software to enable quantitative evaluation of parenchyma enhancement and to describe perfusion variables.25–29 Contrast-enhanced ultrasonography is a minimally invasive technique and applicable in many clinical settings; therefore, it may become the modality of choice for evaluation of renal blood flow in response to a physiologic stimulus and for monitoring during a perfusion deficit.
Given the complex and heterogeneous nature of the renal vasculature, knowledge of the anatomy of the kidneys of dogs is important. The renal artery divides into 2 branches, which in turn divide into 2 to 4 interlobar arteries. These divide into arcuate arteries at the corticomedullary junction. The arcuate arteries radiate toward the periphery of the cortex, where they divide into numerous interlobular arteries. Afferent arterioles arise from the interlobular arteries to supply the glomeruli and then merge to form efferent arterioles. Venous drainage of the kidneys is via the numerous stellate veins in the fibrous portion of the capsule. These connect with veins in the adipose portion of the capsule and empty into interlobular, arcuate, and interlobar veins before entering the main trunk of the renal vein. Venous arcuate vessels, in contrast to their arterial counterparts, unite to form elaborate arches. Arcuate veins span the medulla to join the dorsal and ventral regions of each kidney.30
The authors are aware of no guidelines for the optimal size of ROIs during CEUS evaluation of perfusion in dogs. In the study reported here, results for several small ROIs were compared with those for a larger ROI, all of which were drawn at the same depth in the near field of the renal cortex, obtained for unsedated dogs to test the hypothesis that the location and size of ROIs did not significantly affect mean values of several renal perfusion variables.
Materials and Methods
Animals
A homogeneous group of 12 client-owned adult (1.5 to 2 years old) Labrador Retrievers (8 males and 4 females; mean ± SD body weight, 27 ± 1.6 kg), all of which were fed similar commercial hydrolyzed protein diets, were included in the study. The dogs were referred for CEUS of the gastrointestinal tract because they had digestive tract disorders, including adverse food reactions and eosinophilic enteritis. A complete clinical examination, blood pressure monitoring, CBC, serum biochemical analysis, and urinalysis were performed for each dog. All owners provided consent for participation of their dogs in the study. All treatments, housing, and animal care were in accordance with EU Directive 2010/63/EU for animal experiments.
Ultrasonographic examinations
Examinations with B-mode ultrasonography, Doppler ultrasonography, and CEUS were performed on all dogs. All ultrasonographic examinations were performed by the same investigator (FM) by use of a scannera equipped with contrast-tuned imaging technology.b The dogs were not sedated and were placed in right lateral recumbency. Hair over the ventral and lateral portions of the abdomen was clipped. Alcohol and coupling gel were applied to the skin, and B-mode ultrasonography of the gastrointestinal tract and both kidneys was performed with micro-convex (5.0- to 8.0-MHz) and linear (10- to 12-MHz) transducers. During ultrasonographic examination, both kidneys were measured in the longitudinal plane, and the maximum craniocaudal diameter was determined. Arterial and venous blood flows were visually examined by use of color Doppler ultrasonography. A pulsed-wave recording was performed on the interlobar artery and arcuate artery. Doppler measurements were repeated 3 times, and a mean value was calculated. Renal RI was calculated by use of the following equation:
RI = (peak systolic flow velocity – end diastolic flow velocity)/peak systolic flow velocity
Contrasted-enhanced ultrasonographic examinations were performed immediately following B-mode ultrasonography for all dogs; a linear transducer (5.0- to 7.5-MHz) probe with contrast agent capability was used for CEUS. The contrast agent was a sulfur hexafluoride signal enhancer.c The contrast agent was prepared in accordance with the manufacturer's recommendations. Each vial of contrast agent (which contained 25 mg of freeze-dried powder) was reconstituted by injection of 5 mL of saline (0.9% NaCl) solution; vials then were shaken vigorously for 20 seconds. An aliquot of the reconstituted suspension (0.5 mL/kg; total volume, approx 1.5 mL) was rapidly infused via a 3-way valve and 18- or 20-gauge catheter inserted in a cephalic vein; the syringe was always maintained in a horizontal position for injection. The catheter was then immediately flushed with 5 mL of saline solution.
Each dog received 2 bolus injections of contrast agent, which were standardized and administered by the same investigator (MD). The first bolus was used for evaluation of the gastrointestinal tract. After evaluation of the gastrointestinal tract was completed, a high-energy ultrasound pulse was delivered to destroy residual microbubbles in the gastrointestinal tract and aorta; the pulse was continued until the background echogenicity was similar to that seen before injection of the contrast medium.
The second bolus of contrast agent was injected approximately 5 to 10 minutes after the first bolus injection; it was used for examination of tissues of the left kidney. Injection of the contrast agent and activation of a timer were performed simultaneously. A longitudinal view of the left kidney was obtained, and dynamic changes during enhancement were recorded for 2 minutes. The left kidney was scanned continuously during the early arterial and late corticomedullary phases by use of a mechanical index set at a low value (0.09). Standardized settings were used for depth (4 cm for the renal cortex), time gain compensation, overall gain, and focal zone. Only 1 focal point was used, which was placed immediately below the left kidney image.
Data processing
Raw data (good-quality video clips) obtained during CEUS were stored digitally on a hard disk. All functional data were analyzed by a trained investigator (LL).
For each dog, 4 ROIs were manually drawn in the near field of the renal cortex. Three small oval ROIs (1, 2, and 3, numbered from left to right; area of each ROI, 0.11 cm2) located in a row with a distance of 1 mm between adjacent ROIs and 1 large oval ROI (4; area, 1 cm2) that encompassed the 3 smaller ROIs were drawn in the renal cortex (Figure 1). Depth at which the ROIs were located, which was calculated as the distance between the body wall and left kidney, was approximately 1.5 to 2.0 cm; it was selected on the basis of the depth at which the image had the best homogeneity. During ROI selection, vascular structures and surrounding tissues were excluded. The ROI locations were standardized among the dogs.
Ultrasonogram depicting ROIs in the renal cortex of a dog. The 3 small ROIs (1, 2, and 3 from left to right) are oval, and each has an area of 0.11 cm2. They are located in a row in the near field of the renal cortex at a distance of 0.1 cm between adjacent ROIs. The large oval ROI (4) has an area of 1 cm2 and encompasses the 3 smaller ROIs. The ROIs were manually drawn at a depth of approximately 1.5 to 2.0 cm, which is the depth at which the image had the best homogeneity.
Citation: American Journal of Veterinary Research 77, 8; 10.2460/ajvr.77.8.869
Ultrasonogram depicting ROIs in the renal cortex of a dog. The 3 small ROIs (1, 2, and 3 from left to right) are oval, and each has an area of 0.11 cm2. They are located in a row in the near field of the renal cortex at a distance of 0.1 cm between adjacent ROIs. The large oval ROI (4) has an area of 1 cm2 and encompasses the 3 smaller ROIs. The ROIs were manually drawn at a depth of approximately 1.5 to 2.0 cm, which is the depth at which the image had the best homogeneity.
Citation: American Journal of Veterinary Research 77, 8; 10.2460/ajvr.77.8.869
Ultrasonogram depicting ROIs in the renal cortex of a dog. The 3 small ROIs (1, 2, and 3 from left to right) are oval, and each has an area of 0.11 cm2. They are located in a row in the near field of the renal cortex at a distance of 0.1 cm between adjacent ROIs. The large oval ROI (4) has an area of 1 cm2 and encompasses the 3 smaller ROIs. The ROIs were manually drawn at a depth of approximately 1.5 to 2.0 cm, which is the depth at which the image had the best homogeneity.
Citation: American Journal of Veterinary Research 77, 8; 10.2460/ajvr.77.8.869
Postprocessing analysis was performed by use of image-analysis software.d This software allowed processing of the raw data to enable investigators to examine tissue perfusion in ROIs and to automatically calculate variables. A time-intensity curve, which is a parametric curve of time versus SI, was also generated for each ROI. For each ROI, the software generated the variables peak enhancement, TTP, MTT, and RBF (Figure 2). Peak enhancement was defined as the percentage increase in SI (from 0 to 100 as maximal intensity) reached during transit of the contrast agent at a specific time point. The TTP was defined as the interval until maximum SI of the contrast agent. The MTT was defined as the circulation time of the contrast agent in the examined tissue. The RBF was defined as the ratio between regional blood volume and MTT.
A time-intensity curve created by use of quantitative CEUS data. Time 0 was the start of the injection of contrast medium. Notice the markings indicating the variables peak enhancement, which represents the percentage increase in SI reached during transit of the contrast agent at a specific time point; MTT; and TTP.
Citation: American Journal of Veterinary Research 77, 8; 10.2460/ajvr.77.8.869
A time-intensity curve created by use of quantitative CEUS data. Time 0 was the start of the injection of contrast medium. Notice the markings indicating the variables peak enhancement, which represents the percentage increase in SI reached during transit of the contrast agent at a specific time point; MTT; and TTP.
Citation: American Journal of Veterinary Research 77, 8; 10.2460/ajvr.77.8.869
A time-intensity curve created by use of quantitative CEUS data. Time 0 was the start of the injection of contrast medium. Notice the markings indicating the variables peak enhancement, which represents the percentage increase in SI reached during transit of the contrast agent at a specific time point; MTT; and TTP.
Citation: American Journal of Veterinary Research 77, 8; 10.2460/ajvr.77.8.869
Statistical analysis
A 1-way ANOVA was used to compare the effects of location and size of the ROIs on the perfusion variables derived from time-intensity curves. All data were analyzed by use of general linear modelinge and the following equation: Yij = μ + αi + ϵij, where Y is the dependent variable, μ is the general mean value, α is the independent variable ROI (1, 2, 3, or 4), and ϵij is the error. Values were considered significant at P < 0.05.
Results
Information collected regarding kidney function of the dogs (history of renal disease, clinical findings, and laboratory data) did not reveal any abnormalities. Particularly, clinical data including blood pressure, capillary refill time, heart rate, respiratory rate, and hydration status were within expected limits. Laboratory data, including results for a CBC, renal profile hematologic test (concentrations of creatinine, calcium, sodium, chloride, albumin, BUN, proteins, phosphorus, glucose, and potassium), and urinalysis (urine specific gravity, concentration of urinary proteins, and results for examination of urinary sediment) were also within anticipated limits.
Consistent with the lack of abnormalities in the clinical and laboratory data, B-mode ultrasonographic evaluation of both kidneys revealed no evidence of focal or diffuse abnormalities. The cortex was hyperechoic when compared with the medulla but was hypoechoic when compared with the spleen. The medulla appeared to be separated into several lobulated segments by interlobar vessels and diverticuli. No evidence of focal or diffuse abnormalities in the kidneys was found. Sizes of structures were as anticipated. The arcuate and interlobar arteries could be identified at the corticomedullary junction and within the cortex by use of color Doppler ultrasonography. Mean ± SD renal RI was 0.62 ± 0.05.
Contrast-enhanced ultrasonographic examination was used to measure real-time blood flow and renal perfusion in vivo by use of grayscale imaging after administration of contrast medium. Renal perfusion imaging was performed on the left kidney of each dog. Examination of the image obtained in the longitudinal plane revealed gradual enhancement from the segmental renal arteries to the small interlobular arteries in the renal cortex to the renal medullar arteries.
Contrast enhancement of the kidney was subjectively evaluated by 1 investigator (FM). In the inflow phase, there was an initial increase in echogenicity of the cortical area, whereas the medulla had slower enhancement. The renal medulla appeared isoechoic when compared with the cortex at approximately 30 seconds after injection of the contrast medium (Figure 3). In the outflow phase, there was a gradual and homogeneous decrease of enhancement, which was more intense in the renal cortex than in the medulla.
The CEUS images of the vascularization pattern in the left kidney of a representative dog obtained before (A) and 15, 30, and 60 seconds after (B, C, and D, respectively) injection of a bolus of contrast medium. In panel D, notice that the contrast agent is distributed homogenously within the kidney and that the medulla appears isoechoic relative to the renal cortex. Tick marks at the bottom of each image are at intervals of 1 cm.
Citation: American Journal of Veterinary Research 77, 8; 10.2460/ajvr.77.8.869
The CEUS images of the vascularization pattern in the left kidney of a representative dog obtained before (A) and 15, 30, and 60 seconds after (B, C, and D, respectively) injection of a bolus of contrast medium. In panel D, notice that the contrast agent is distributed homogenously within the kidney and that the medulla appears isoechoic relative to the renal cortex. Tick marks at the bottom of each image are at intervals of 1 cm.
Citation: American Journal of Veterinary Research 77, 8; 10.2460/ajvr.77.8.869
The CEUS images of the vascularization pattern in the left kidney of a representative dog obtained before (A) and 15, 30, and 60 seconds after (B, C, and D, respectively) injection of a bolus of contrast medium. In panel D, notice that the contrast agent is distributed homogenously within the kidney and that the medulla appears isoechoic relative to the renal cortex. Tick marks at the bottom of each image are at intervals of 1 cm.
Citation: American Journal of Veterinary Research 77, 8; 10.2460/ajvr.77.8.869
The quantitative analysis software converted the video intensity data into time-intensity curves by fitting the percentage SI to the parametric graph with the γ variate function that best represented the pattern (Figure 4). The ascending segment of the curve was steep; the curve then gradually descended to the zero value. Intersection of the cortical and medullary time-intensity curves corresponded to the point of the isoenhancement pattern of renal parenchyma (Figure 5).
Time-intensity curves created by use of CEUS images of the left kidney of a representative dog obtained after injection of contrast agent. Perfusion software was used to analyze the data. The jagged gray line was generated from actual data, whereas the curved black line was generated from data fitted by use of a corrected γ function. Values determined for perfusion variables for the curved line were as follows: peak enhancement, 29.8%; MTT, 45.64 seconds; TTP, 24.245 seconds; and RBF, 39.440 L/min.
Citation: American Journal of Veterinary Research 77, 8; 10.2460/ajvr.77.8.869
Time-intensity curves created by use of CEUS images of the left kidney of a representative dog obtained after injection of contrast agent. Perfusion software was used to analyze the data. The jagged gray line was generated from actual data, whereas the curved black line was generated from data fitted by use of a corrected γ function. Values determined for perfusion variables for the curved line were as follows: peak enhancement, 29.8%; MTT, 45.64 seconds; TTP, 24.245 seconds; and RBF, 39.440 L/min.
Citation: American Journal of Veterinary Research 77, 8; 10.2460/ajvr.77.8.869
Time-intensity curves created by use of CEUS images of the left kidney of a representative dog obtained after injection of contrast agent. Perfusion software was used to analyze the data. The jagged gray line was generated from actual data, whereas the curved black line was generated from data fitted by use of a corrected γ function. Values determined for perfusion variables for the curved line were as follows: peak enhancement, 29.8%; MTT, 45.64 seconds; TTP, 24.245 seconds; and RBF, 39.440 L/min.
Citation: American Journal of Veterinary Research 77, 8; 10.2460/ajvr.77.8.869
Time-intensity curves created by use of CEUS images of the left kidney of a representative dog. The intersection of the curves for the cortex (dotted line) and medulla (solid line) corresponds to the point of the isoenhancement pattern of renal parenchyma.
Citation: American Journal of Veterinary Research 77, 8; 10.2460/ajvr.77.8.869
Time-intensity curves created by use of CEUS images of the left kidney of a representative dog. The intersection of the curves for the cortex (dotted line) and medulla (solid line) corresponds to the point of the isoenhancement pattern of renal parenchyma.
Citation: American Journal of Veterinary Research 77, 8; 10.2460/ajvr.77.8.869
Time-intensity curves created by use of CEUS images of the left kidney of a representative dog. The intersection of the curves for the cortex (dotted line) and medulla (solid line) corresponds to the point of the isoenhancement pattern of renal parenchyma.
Citation: American Journal of Veterinary Research 77, 8; 10.2460/ajvr.77.8.869
The CEUS quantitative variables did not differ among the 4 ROIs drawn at the same depth in the near field of the renal cortex where the image had the best homogeneity. Statistical analysis of the CEUS-derived data revealed no significant differences in mean values of renal perfusion variables (peak enhancement, TTP, RBF, or MTT) associated with the location or size of each ROI (Table 1).
Mean ± SD values for perfusion variables determined by use of ROIs* drawn manually on the CEUS image of the cortex of the left kidney of each of 12 dogs.
| Variable | ROI 1 | ROI 2 | ROI 3 | ROI 4 | P value† |
|---|---|---|---|---|---|
| Peak enhancement (%)‡ | 17.16 ± 6.89 | 19.79 ± 6.27 | 17.23 ± 8.31 | 18.06 ± 7.73 | 0.37 |
| TTP (s) | 30.18 ± 9.86 | 23.46 ± 7.36 | 25.74 ± 7.59 | 25.46 ± 5.66 | 0.08 |
| RBF (L/min)§ | 19.51 ± 9.02 | 24.37 ± 9.03 | 20.44 ± 10.28 | 21.47 ± 9.43 | 0.16 |
| MTT (s) | 43.42 ± 17.04 | 38.75 ± 13.11 | 37.16 ± 6.84 | 39.14 ± 10.19 | 0.18 |
The 3 small ROIs (1, 2, and 3) were oval, and each had an area of 0.11 cm2. They were located in a row in the near field of the renal cortex at a distance of 0.1 cm between adjacent ROIs. The large oval ROI (4) had an area of 1 cm2 and encompassed the 3 smaller ROIs. The ROIs were manually drawn at a depth of approximately 1.5 to 2.0 cm, which was the depth at which the image had the best homogeneity.
Values were considered significant at P < 0.05.
Peak enhancement was defined as the percentage increase in SI reached during transit of the contrast agent at a specific time point.
Calculated as the ratio between regional blood volume and MTT.
Discussion
The development of CEUS with microbubble contrast agents has provided a unique means of visually evaluating and quantifying tissue perfusion, which offers substantial advantages in terms of real-time imaging, cost, and patient safety.31 Quantitation of tissue perfusion represents one of the most promising areas for use of CEUS. In particular, ultrasonographic contrast agents will enable the development of new functional applications for renal blood flow quantification in evaluation of renal function.28
Clinical and laboratory data and urinalysis results allowed us to verify that the dogs in the study reported here did not have clinical evidence of renal failure in the form of subclinical or progressive renal disease. Renal vascular resistance was evaluated by measuring RI as described elsewhere.32–35 The mean RI value was within the reference range for dogs,36,37 which confirmed no alterations in renal blood flow.
Several factors may contribute to variability in quantitative analysis of renal CEUS and the resulting perfusion variables. These factors can be divided into 3 categories: factors related to technical variables, including scanner settings, transmission power, focal zone, dynamic range, gain setting, time gain compensation, and transmission frequency; factors related to the contrast medium, including type, preparation, injection technique, and dose; and patient-related factors, including physiologic differences (heart rate, blood pressure, and respiratory rate), physiologic interactions of the patient with the agent microbubbles, variability of propagation and attenuation of ultrasonographic waves in tissues, and hemodynamic variability among species.9,31–41 To minimize the influence of these factors, the methods used in the present study were standardized as much as possible (eg, imaging settings were consistent among all dogs, and the same investigators performed the procedures on all dogs). A low mechanical index (0.09) was chosen to minimize microbubble disruption and allow accumulation of microbubbles in the microvasculature,26 which has been suggested to avoid variability in clinical applications of CEUS.31 Time gain compensation and overall gain were decreased before injection of the contrast agent to suppress most background tissue signals.
The type of contrast agent can affect results of quantitative imaging analysis. In addition, the preparation, dose, method used during injection, and effects of the handling methods on the microbubbles can influence the physical properties and acoustic behavior of the agent microbubbles during quantitative imaging. All the commercial agents have broad distributions for microbubble size, and the mean size differs substantially among agents, which makes it difficult to compare quantification analysis performed with different agents.31 Furthermore, for the same contrast agent, the manner in which it is reconstituted prior to administration may introduce substantial variation in both size and concentration of microbubbles. In the present study, a sulfur hexafluoride echo-signal enhancer was used; the manufacturer's recommendation was reconstitution through the addition of saline solution followed by vigorous manual shaking. To standardize the preparation, this shaking step was performed by the same investigator for all vials of contrast agent. Other factors that were controlled because they may have influenced the interval until peak enhancement included use of a 3-way valve, consistency in the volume of saline solution used to flush the catheter, and rapidity of contrast agent administration.26
The dogs of the present study were considered medium to large in size; thus, an IV catheter with a rather large needle diameter (18 gauge or 20 gauge) was used to avoid reduction of the concentration of microbubbles.31 The contrast agent was administered as a bolus injection into a cephalic vein by use of a 3-way valve; the syringe was always maintained in a horizontal position for injection. The dose of the contrast medium was the same for each dog, and a constant volume of saline solution was used to flush the catheter immediately after injection of the contrast agent. Thus, we attempted to minimize the effects of the handling of the microbubbles on results of the quantitative imaging analysis. The data analysis was also standardized by use of software that could be useful for standardizing and reproducing quantitative CEUS evaluations.
Patient-related factors can have a great impact on the perfusion data obtained.31 Cardiac output (affected by stress, age, sex, breed, and size) may influence the interval until peak enhancement and MTT.17 Moreover, the size of the organ or tissue may influence the acoustic power in the ultrasonographic field (power is reduced along the sides and in the far field) and may cause variations in SI of the peak.17
The group of dogs included in the present study was as homogeneous as possible. The dogs were of the same breed, had uniform age and body weight, and had clinical and laboratory data that indicated no abnormalities in renal function. Blood pressure was within reference limits for all dogs. To avoid effects of blood pressure on peak intensity, ROIs were always drawn in an area located distant from large arteries, as has been suggested in another study.31
Some sedation and anesthesia protocols for dogs and cats may influence contrast medium dynamics through their effects on the cardiovascular system.19,22 It is interesting that in a study42 on sedation protocols for feline patients to enable investigators to perform renal CEUS evaluations, butorphanol had favorable cardiovascular properties without significant effects on renal perfusion variables. In the present study, the dogs were not sedated or anesthetized during CEUS. Therefore, possible effects on renal blood flow attributable to these drugs can be ruled out for this study.
There are conflicting data regarding the optimal method for selecting an ROI in quantitative CEUS analysis of renal perfusion. Some authors recommend that the ROI should enclose the largest area of visible renal cortex to minimize the influence of local perfusion heterogeneities.38,39 In contrast, other researchers have reported a significant inverse association between the size of ROI and the peak intensity.24 In the present study, 3 small ROIs with the same size and shape were drawn at the same depth of the renal cortex, as was reported in another study.24 Comparison of results for the smaller ROIs with results for the large ROI was performed as reported elsewhere.39 Investigators in that study39 chose to draw the largest ROI possible. In accordance with these previous reports, we were extremely careful to avoid the inclusion of vascular structures within an ROI, which could have affected the perfusion data. Furthermore, the 3 small ROIs were positioned at locations in the renal cortex that could be easily repeated in all dogs.
Consistent with results of another study,24 results for the study reported here indicated no significant differences among the perfusion variables for the ROIs located in a row in the near field of the renal cortex. Results for CEUS in the present study also indicated that there were no significant differences between the mean values of the renal perfusion variables (peak enhancement, TTP, RBF, and MTT) for each small ROI (0.11 cm2), compared with the value for the large ROI (1 cm2). This likely depended on the accurate selection of the ROIs and on the use of the selected software. Avoiding errors in SI analysis caused by tissues (eg, vessels) that should not be included in an ROI is dependent on the use of software that allows frame-by-frame editing to eliminate erroneous frames (ie, frames with abnormal intensities or spikes) and that makes corrections for involuntary movements (eg, breathing).
The 3 smaller ROIs were inside the large ROI. Thus, heterogeneity of diffusion of the contrast medium in the area was eliminated. This resolved a weakness of another study24 in which the ROIs were drawn separately and not inside one another. This can be crucial for explaining the difference in peak intensity values.
Qualitative analysis of the CEUS data during the present study indicated that the renal medulla had the same enhancement as the renal cortex at approximately 30 seconds after injection of the contrast medium. Examination of the corresponding time-intensity curves confirmed this finding, as indicated by the intersection of the curves. This result is in agreement with the results of other studies9,40 in which a sulfur hexafluoride contrast agent was used to study renal perfusion in dogs. In particular, investigators of 1 study40 reported a consistent homogeneity of the renal parenchyma at approximately 30 seconds after injection of the contrast medium. Conversely, investigators of another study26 used a perfluoropropane contrast medium and found that the medulla remained hypoechoic when compared with the cortex throughout contrast medium inflow and outflow periods. We believe this may have been attributable to the differences in enhancement behavior for the 2 contrast agents.
In the present study, a longer interval to peak perfusion of the renal cortex was found, compared with the value reported in another study.26 However, this finding did not affect comparison among the ROIs. This finding could have been related to the type of contrast agent used or to the breed of the dogs. The dogs in the present study had gastrointestinal disorders, although they had no abnormalities of renal function. It cannot be excluded that these gastrointestinal disorders may have affected transit time of the contrast medium within the renal parenchyma. Additional CEUS examinations could be performed to investigate renal blood flow variables in healthy dogs and changes related to the inclusion of other breeds of dogs.
Analysis of renal perfusion through quantitative evaluation of enhancement with a contrast agent into a large enough area of renal cortex may be of clinical importance in the diagnosis of diffuse renal failure. Results of the present study could be useful in defining guidelines for the selection of ROIs and controlling variability in the use of CEUS to evaluate renal perfusion. In patients with chronic renal disease, impairment of perfusion is an early event in the course of renal dysfunction and usually precedes functional damage43; therefore, quantitative CEUS analysis may be used for early evaluation of renal dysfunction44 to facilitate the management of this pathological process.
Acknowledgments
No third-party funding or support was received in connection with this study or the writing or publication of the manuscript. The authors declare that there were no conflicts of interest.
ABBREVIATIONS
| CEUS | Contrast-enhanced ultrasonography |
| MTT | Mean transit time |
| RBF | Regional blood flow |
| RI | Resistive index |
| ROI | Region of interest |
| SI | Signal intensity |
| TTP | Time to peak |
Footnotes
MyLab 40/Vet, Esaote, Genova, Italy.
CnTI, Esaote, Genova, Italy.
SonoVue, Bracco Imaging, Milan, Italy.
Qontrast, Bracco Imaging, Milan, Italy.
PROC GLM, SAS, version 8.1, SAS Institute Inc, Cary, NC.
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