Ultrasonography of Osteosarcoma in Dogs
Sonography Atlas of Dogs

Abstract

Heart rate and blood pressure monitoring: The heart rate (HR) and indirect blood pressure [(BP): systolic arterial pressure (SAP), mean arterial pressure (MAP) and diastolic arterial pressure (DAP)] were measured and recorded at time 0 (before anesthetics injection) and at 5, 10 and 20 min after anesthetics administration using a patient monitor (VP-700^®, Votem, Chuncheon, Korea) and oscillometric BP device (VET HDO monitor^®, Vetline LCC Co., Babenhausen, Germany).

Statistical analysis: Repeated ANOVA was used to compare PI, upslope, downslope, TTP₀, TTP_up and AUC among the three groups of dogs. Bonferroni correction was applied to account for multiple comparisons. Repeated ANOVA was also used for comparison of HR and BP between the three groups with Bonferroni correction to account for multiple comparisons. For all analyses, P<0.05 was considered statistically significant. All statistics were performed using a computer statistical package (IBM SPSS Statistics for Windows, version 20.0, IBM Corp., Armonk, NY, U.S.A.).

RESULTS

Real-time observation of renal perfusion: The contrast ultrasound techniques were easy to perform, and contrast enhancement was clearly visible in both conscious, and anesthetized dogs. In conscious dogs, there was rapid, homogeneous contrast enhancement throughout the renal cortex; this enhancement was subjectively more hyperechoic than in the renal medulla. The cortex had a rapid initial inflow of contrast enhancement followed by a fade-out during which the medulla would begin to enhance. The initial contrast enhancement of the medulla was invariably more delayed than in the renal cortex and usually remained coarser and hypoechoic compared with the renal cortex in all dogs. The enhancement patterns of the cortex and medulla are displayed in Fig. 2. Subjectively, in anesthetized dogs (TZM and M groups), the enhancement pattern was similar to that of the conscious group. The enhancement wash-in was delayed. However, the degree of PI of the cortex and medulla, and the heterogeneity of contrast enhancement among the three groups did not differ. These observations are supported by objective results (Tables 1 andand 2Table 2).

Figure 2: Contrast enhanced images of the left kidney in different stages after a bolus injection of contrast agent. (A) 6 sec after the injection; base line; (B) 9 sec: the contrast agent is diffusely spread in the cortex; (C) 11 sec: the cortical enhancement has reached its peak; the medulla is not enhanced; (D) 15 sec: the contrast agent in the cortex is starting to fade; (E) 22 sec: the medullary perfusion is at its peak; (F) 70 sec: the contrast agent is fully washed out of the cortex and medulla.

Table 2: Mean and standard deviation of renal cortical perfusion parameters in dogsData are expressed as mean ± SD (n=8). TZM, tiletamine-zolazepam-medetomidine; M, medetomidine; TTP₀, time to peak enhancement from contrast injection; TTP_up, time to peak enhancement from initial rise; AUC, area under the curve. a) P<0.05 (Conscious group versus M group); b) P<0.05 (Conscious group versus TZM group); c) P<0.05 (TZM group versus M group).

Table 2: Mean and standard deviation of renal medullary perfusion parameters in dogsData are expressed as mean ± SD (n=8). TZM, tiletamine-zolazepam-medetomidine; M, medetomidine; TTP₀, time to peak enhancement from contrast injection; TTP_up, time to peak enhancement from initial rise; AUC, area under the curve. a) P<0.05 (Conscious group versus M group); b) P<0.05 (Conscious group versus TZM group); c) P<0.05 (TZM group versus M group).

Quantitative analysis of renal perfusion: Representative time-intensity curves from each group are shown in Fig. 3. Compared to the cortical perfusion parameters in the conscious group, TTP₀ was significantly delayed in the TZM group, while the upslope, TTP₀ and TTP_up differed significantly in the M group (P<0.05). The AUC of the cortex in the TZM and M groups was not different with that of the conscious group (Table 1). Analysis of renal medullary parameters (Table 2) showed significantly decreased upslope in the TZM and M groups (P<0.05). TTP₀ and TTP_up were significantly delayed in the TZM and M groups (P<0.05). The AUC of the medulla was significantly decreased in the M group (P<0.05) compared with the concious and TZM groups. The upslope, TTP₀ and TTP_up of the M group were significantly different from those of the TZM group (P<0.05).

Figure 3: Time-intensity curves obtained from three representative dogs from the A) conscious group, B) TZM group and C) M group.

HR and BP: Data related to HR, SAP, MAP and DAP are shown in Fig. 4. There were no significant differences in SAP, MAP and DAP between anesthetized and conscious dogs. However, in the M group, the HR was significantly lower 10 min and 20 min after administration of medetomidine compared to the HR before anesthetic injection (P<0.0083).

Figure 4: Blood pressure and heart rate following administration of anesthetics in the tiletamine-zolazepam and medetomidine (TZM) and metetomidine (M) groups.

DISCUSSION

In this study, the contrast enhancement pattern (Fig. 3) of the conscious group was similar to that in previous CEUS reports of the normal canine kidneys [13, 21]. However, intensity parameters are difficult to compare with the previous results, because intensity is easily affected by variable machine settings, such as gain, transducer frequency and the mechanical index. However, these factors do not affect the slope or time to peak [21]. In the present study, the mean time to peak was 10.33 ± 1.73 sec in the renal cortex and 16.56 ± 2.69 sec in the renal medulla. One study on CEUS of kidneys in eight dogs showed relatively delayed mean time to peak compared with our results. This discrepancy was thought to be due to the differences in the speed and volume of contrast agent injection, volume of saline flush and patient factors like cardiac output or blood pressure, which can be affected by anesthetic protocols. In addition, body weight can cause the discrepancy of the results. Our dogs were smaller than those in the previous study (8–25 kg). In a CEUS study on normal canine spleens, the association between body weight and the perfusion variables was evaluated between small dogs (≤15 kg) and larger dogs (≥15 kg). The time to peak intensity and AUC were lower in small dogs, indicating overall perfusion is lower and the rate of inflow is faster than those in larger dogs [16]. However, their results were not statistically significant. Therefore, renal perfusion variables should be evaluated with respect to body weight.

Both the size and location of the ROI can have an effect on the echo-intensity value. The larger or deeper the ROI, the lower the PI [13]. ROI size is recommended to be large enough to represent the tissue being analyzed, but small enough to avoid the surrounding structure and vessels, and the ROI placement for perfusion quantification should be where enhancement is the most homogeneous and close to the focal zone [13]. Therefore, in this study, the size of the ROI was designated as 3 × 3 mm², which was large enough to include the parenchyma of the kidney without interference from large vessels or respiratory motion. The depth of the ROI was set between 1 and 1.5 cm to avoid attenuation of tissue and analyze the most homogeneous part of the kidney parenchyma. In this study, standardization of the ROI size and location resulted in PI values of the cortex and medulla that did not differ between the groups.

Compared to the conscious group, the M group had a significantly delayed TTP₀ and TTP_up of the cortex and medulla. These results might be caused by decreased cardiac output and HR after medetomidine administration [3]. In this study, cardiac output was not measured, but likely decreased. The HR decreased significantly after 10 min following administration of medetomidine in the M group (Fig. 4). Decreased cardiac output and HR affect the renal blood flow; therefore, the slowly infused contrast agent reaching the kidneys resulted in delayed TTP₀ and TTP_up. Because the upslope value was calculated based on TTP_up and an increase in echo-intensity, the delayed TTP_up primarily affected the upslope values of the renal cortex and medulla. The effect of medetomidine can be rapidly reversed with a specific alpha 2-adrenoceptor antagonist, atipamezole. This is an advantage for simple procedure like CEUS [3]. However, the side effects of medetomidine caused marked bradycardia, and hence, decreased cardiac output resulted in remarkable changes in renal perfusion parameters during CEUS in dogs receiving only medetomidine in this study.

By minimizing the dose of medetomidine in the TZM group, HR and BP were maintained during anesthesia. The blood flow rate to the kidney seemed to be minimally affected in the TZM group. Therefore, there were no changes in the perfusion parameters, except for the TTP₀ in the cortex. Although the combination of anesthetics was different from the present study, a previous report described that a medetomidine-midazolam-butorphanol combination decreases intestinal and skeletal muscle microvascular blood flows, while renal cortical blood flow is maintained in dogs [17]. In that study, renal medullary flow was not examined. In another study on the effects of iatrogenic hypercortisolism on kidneys in dogs, renal medulla showed similar changes with those of cortex, which reflect that medullary blood flow depends on changes in cortical flow [6]. However, renal cortical and medullary blood flows have been reported to react differently to arterial chemoreceptor stimulation anesthetics [11]. When renal blood perfusion parameters in the renal medulla were compared with those of the cortex in the present study, TTP_up and upslope also changed under TZM anesthesia. These results are presumably because this anesthetic combination induced different reactions in the renal cortex and medulla. PI, AUC and down slope in the renal medulla did not show significant changes like those of the cortex, which suggests that renal medullary blood volume was maintained under TZM anesthesia.

Our study has several limitations. Firstly, we examined only the left kidney (left kidney), because it is impossible to perform CEUS on both kidneys simultaneously. A study that evaluated glomerular filtration rate with dynamic computed tomography in normal dogs reported that contrast agent concentrations and glomerular filtration rate were not significantly different between the left and right kidneys [2]. Therefore, it is thought that the perfusion of one kidney is similar to the other. Secondly, 24-gauge IV catheters were used in this experiment, and injections were performed manually. These techniques can cause possible microbubble destruction and injection time variability. In a study of the effect of needle size and injection rate on microbubble contrast agent population, a 23-guage needle was relatively nondestructive to microbubble contrast agents [20]. The authors reported needles smaller than 27-gauge and rapid injection rate to be the major factors associated with microbubble destruction. Therefore, usage of 24-gauge needles should not have caused significant microbubble destruction, however, larger catheters should have been used in this experiment. Thirdly, inter-observer and intraobserver variability was not evaluated in this study, although CEUS examinations are known to be examiner-independent. Lastly, necropsy and histopathologic confirmation of the kidney condition were not performed in all dogs in this study.

In conclusion, renal cortical perfusion pattern under TZM anesthesia was not significantly different with conscious status, except for TTP₀. However, perfusion factors in the renal medulla were affected by anesthesia during CEUS. Therefore, the TZM combination may be a useful anesthetic protocol for evaluating renal cortical perfusion, and data obtained in this study may serve as reference values for evaluation of renal perfusion using CEUS in dogs under the injectable anesthesia

Acknowledgments

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