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Influence of ageing on quantitative contrast-enhanced ultrasound of the kidneys in healthy cats
  1. Emmelie Stock1,
  2. Dominique Paepe2,
  3. Sylvie Daminet2,
  4. Luc Duchateau3,
  5. Jimmy H Saunders1 and
  6. Katrien Vanderperren1
  1. 1 Department of Medical Imaging and Small Animal Orthopaedics, Faculty of Veterinary Medicine, Ghent University, Merelbeke, Belgium
  2. 2 Small Animal Department, Faculty of Veterinary Medicine, Ghent University, Merelbeke, Belgium
  3. 3 Department of Comparative Physiology and Biometrics, Faculty of Veterinary Medicine, Ghent University, Merelbeke, Belgium
  1. E-mail for correspondence; Emmelie.Stock{at}UGent.be

Abstract

The degenerative effects of ageing on the kidneys have been extensively studied in humans. However, only recently interest has been focused on renal ageing in veterinary medicine. Contrast-enhanced ultrasound allows non-invasive evaluation of renal perfusion in conscious cats. Renal perfusion parameters were obtained in 43 healthy cats aged 1–16 years old, and the cats were divided in four age categories: 1–3 years, 3–6 years, 6–10 years and over 10 years. Routine renal parameters as serum creatinine, serum urea, urine-specific gravity, urinary protein:creatinine ratio and systolic blood pressure were also measured. No significant differences in any of the perfusion parameters were observed among the different age categories. A trend towards a lower peak enhancement and wash-in area under the curve with increasing age, suggestive for a lower blood volume, was detected when comparing the cats over 10 years old with the cats of 1–3 years old. Additionally, no significant age-effect was observed for the serum and urine parameters, whereas a higher blood pressure was observed in healthy cats over 10 years old.

  • ageing
  • renal perfusion
  • contrast-enhanced ultrasound
  • cat
  • kidneys
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Introduction

Renal ageing has been a hot topic in human research for several years, whereas it has only recently gained interest in veterinary medicine.1 2 The population of senior and geriatric cats has been increasing in the past years.3–5 Consequently, effects of ageing on animal health have grown more interest and more attention is paid to health screening in elderly cats.6

In people, ageing is associated with structural changes accompanied by progressive deterioration of renal function. A reduction in renal mass and glomerular number is present in elderly people, paralleled by an increased percentage of sclerotic glomeruli.2 Moreover, glomerular filtration rate (GFR) and renal plasma flow (RPF) decrease with age, whereas renal vascular resistance increases.2 7 8 The reduction in renal blood flow is thought to be primary and precede the changes in GFR and renal morphology.8 This decline appears to be a part of the normal physiological process of cellular and organ senescence.2 Information on the influence of ageing on renal morphology and function in veterinary medicine is limited. An age-related decrease in GFR has been described in small breed dogs, although no significant age-effect on GFR was present for cats.9 In one study, a lower GFR was present in older cats compared with young cats as measured with the exogenous creatinine clearance test; however, no age-effect was present when GFR was measured with exo-iohexol, endo-iohexol and chromium-51 EDTA.10 A higher serum creatinine concentration (sCr) has been found in elderly cats.6 9 11

Contrast-enhanced ultrasound (CEUS) is a functional imaging technique that allows evaluation of tissue perfusion through the use of intravenously injected contrast agent. The ultrasound contrast agent is composed of millions of microbubbles which are tiny gas-filled spheres, stabilised by an outer shell. These microbubbles have a size similar to those of red blood cells, consequently they do not pass the endothelium and remain strictly intravascular.12 13 Promising results have been obtained with CEUS for the evaluation of diffuse renal disorders in both human and veterinary medicine.14–18

The objective of this study was to determine the effect of age on several perfusion parameters obtained with CEUS of the kidneys in healthy cats divided in four age groups. The hypothesis was that ageing would be associated with a decline in renal blood volume and a higher vascular resistance seen as a decrease in slope of the time-intensity curve.

Materials and methods

Healthy cats of ≥1 year were included. The cats were divided into four age groups: 1–3 years, 3–6 years, 6–10 years and over 10 years. Ten to 12 cats were included in each group. Cats were considered healthy if no significant abnormalities were reported in the history, nor detected on physical examination, complete blood count, serum biochemistry profile, thoracic radiographs and abdominal ultrasound. Cats with both sCr >161.8 µmol/mL11 and urine-specific gravity <1.035 were excluded. Only cats free of any medication for at least two months were included.

All owners were informed about the study and gave their written consent.

A routine physical examination, including non-invasive measurement of systolic blood pressure (SBP) by Doppler ultrasonic technique following the consensus statement of the American College of Veterinary Internal Medicine,19 was performed in all cats. Thyroid palpation was performed in all cats ≥6 years. Five mL of blood was collected by jugular venipuncture. Blood work consisted of complete blood count and serum biochemistry, including total thyroxine (TT4) level in cats of ≥6 years. Five to 10 mL of urine was collected by ultrasound-guided cystocenthesis. Urinalysis included sediment analysis as previously described,6 specific gravity (USG), urinary protein:creatinine ratio (UPC), dipstick and urine culture. Thoracic radiographs (left-right lateral and ventrodorsal projections) and complete abdominal ultrasound were performed.

For the administration of ultrasound contrast agent, a 22-gauge indwelling catheter was placed in the cephalic vein. The hair was clipped over the ventrolateral aspect of the abdomen and coupling gel was applied to the skin. The ultrasound examinations were performed unsedated with the cat manually restrained in dorsal recumbency. The kidney of interest was centred on the screen and was imaged in a longitudinal plane using dual-screen (simultaneous display of conventional B-mode and contrast-mode image). The transducer was manually positioned during each imaging procedure and was maintained at the same position during the CEUS examination.

The contrast agent (Sonovue, Bracco, Italy), 0.05 mL/kg, was injected intravenously (bolus injection over ±3 s) followed by injection of 1.5 mL saline bolus. A three-way stopcock was used to avoid any delay between the injection of contrast agent and saline. The same person performed the injection in a standardised way. Three injections of contrast were performed: two for the left kidney and one for the right kidney. Only the second and third injections were used for further analysis.20 Between subsequent injections, to avoid artefacts, remnant microbubbles were destroyed by setting the acoustic power at the highest level and scanning the caudal aspect of the abdominal aorta for approximately two minutes.

All examinations were performed using a linear transducer of 12–5 MHz on a dedicated machine (iU22, Philips) with contrast-specific software. Basic technical parameters were a single focus placed under the kidney, persistency off, mechanical index 0.09, high dynamic range setting (C50), timer started at the beginning of the injection, gain 85 per cent (corresponding to a nearly dark/anechoic image before USCA administration). The settings were standardised. All studies were digitally registered as a movie clip at a rate of nine frames per second, during 90 seconds.

The clips were exported in DICOM format and analysed off-line using specialised computer software (VueBox, Bracco Suisse SA, Geneva, Switzerland) for objective quantitative analysis. Six regions-of-interest (ROIs) were manually drawn by the same person: three in the renal cortex, two in the renal medulla and one on an interlobar artery. The ROIs were similar in size and drawn at the same depth for every region. The placement of the ROIs is illustrated in Fig 1. For every ROI, the software determined mean pixel intensity as a function of time creating a time-intensity curve. Time-intensity curves were analysed for peak enhancement (PE), wash-in area under the curve (WiAUC), rise time (RT), mean transit time (mTT), time to peak (TTP), wash-in rate (WiR), wash-in perfusion index (WiPI; WiAUC/RT), wash-out area under the curve (WoAUC), total area under the curve (AUC), fall time (FT) and wash-out rate (WoR). The values for the three ROIs in the renal cortex and two ROIs in the renal medulla were averaged. Peak enhancement, for the cortex and medulla were normalised to the values obtained for the interlobar artery.

FIG 1:

Contrast-enhanced ultrasound image of the right kidney of a healthy cat obtained at peak enhancement, illustrating placement of the regions-of-interest (ROIs): three ROIs are present in the renal cortex (green), two in the medulla (orange) and one at an interlobar artery (purple).

Statistical analyses were performed with SAS (SAS V.9.4, SAS Institute). Perfusion parameters obtained with CEUS, sCr, serum urea, USG, UPC and SBP were compared among the different age groups, using a linear fixed effects model with age as categorical fixed effect. A global significance level of 5 per cent was used and adjusted P values were calculated for pairwise comparisons.

Results

Forty-nine cats were recruited, however six cats were excluded: two cats based on abnormalities on thoracic x-rays (severe bronchial pattern in both cats with additional pulmonary mass in one cat), severe jejunal wall thickening and intestinal lymphadenopathy in one cat and uncooperative behaviour in three cats. Finally, 43 cats entered the study: 41 domestic shorthaired or longhaired cats, and 2 Ragdoll cats. The mean±sd age of the total population was 6.5±3.9 years, with 10 cats between 1 and 3 years (mean: 1.8 years, median: 1.5 years, range: 1–2.5 years), 11 cats between 3 and 6 years (mean: 4.5 years, median: 4.8 years, range: 3.1–5.8 years), 12 cats between 6 and 10 years (mean: 7.3 years, median: 7.2 years, range: 6.3–8.3 years) and 10 cats older than 10 years (mean: 12.4 years, median: 12.3 years, range: 10–16.5 years). The mean±sd body weight was 4.0±0.7 kg, with the body condition score ranging from 4/9 to 8/9. No significant abnormalities were seen on thoracic x-rays and routine abdominal ultrasound. The kidneys of all included cats were normal in size (3.0–4.6 cm), had a smooth, regular outline and a normal internal architecture.21

SBP, sCr, serum urea, USG and UPC are summarised in Table 1. No significant differences for sCr, serum urea, USG and UPC were observed among the different age groups. A significantly higher SBP was present for the cats between 3 and 6 years compared with the cats between 1 and 3 years (P=0.043), and for the cats over 10 years compared with the cats between 1 and 3 years (P=0.005).

TABLE 1:

Baseline characteristics for the included cats divided into four age groups

No significant changes in renal perfusion parameters with age were noticed in this study. However, a tendency towards a lower PE (P=0.078) and WiAUC (P=0.078) for the renal cortex were present when comparing the cats between 1 and 3 years old and the cats over 10 years old.

The CEUS parameters for the renal cortex and medulla are summarised in Table 2. Fig 2 illustrates the normal perfusion pattern of the kidney in healthy cats.

TABLE 2:

Perfusion parameters obtained with CEUS for the cats divided into four age groups. The data are expressed as mean±sd

FIG 2:

Sequential images demonstrating the normal perfusion pattern obtained with contrast-enhanced ultrasound of the kidney in healthy cats. Early enhancement of the interlobar arteries (5 s), followed by peak cortical enhancement (7 s) and a late corticomedullary phase (20 s).

Discussion

To our knowledge, this is the first study assessing the age-effect on renal perfusion in healthy cats.

No significant changes in renal perfusion parameters were noticed in this study. However, a potentially lower PE and WiAUC was present in elderly cats, indicating a decrease in renal blood volume with age. This corresponds to the findings in human studies, where a reduction in renal blood flow is considered to be a typical functional finding with increasing age.2 7 8 22 Moreover, the renal blood flow reduction in elderly people is thought to be primary and leading to a reduction in GFR and morphological renal changes.8 In humans, age-related changes involve mainly the renal cortex, whereas the medulla is usually unaffected.8 Typical morphological changes in elderly people are increased glomerular sclerosis, reduction in glomerular volume, decrease area and hyalinisation of afferent arterioles and tubulointerstitial fibrosis.2 7 8 23 Similarly in ageing dogs, glomerulosclerosis, interstitial fibrosis and lipofuscin accumulation have been described.1 24 In contrast, no vascular morphological changes were present in aged beagles.24 In dogs, structural changes in ageing kidneys may be clinically silent and not associated with functional deterioration as reported in humans.1 A lower GFR was detected in geriatric small breed dogs, whereas no decline was seen in ageing large breed dogs.9 Variable results are found for sCr in dogs; however, no clinical significant changes were noticed as sCr remained within normal reference intervals in all dogs.9 25 No age-related differences in several urinary markers as urinary albumin:creatinine ratio, urinary C-reactive protein:creatinine ratio, urinary retinol binding protein:creatinine ratio and urinary N-acetyl-beta-D-glucosaminidase:creatinie ratio are reported in healthy dogs.26 Research in ageing cats is even more limited. An age-effect on GFR was only reported in one study using an exogenous creatinine clearance test, whereas no age-effect was seen in other studies.9 10 Higher sCr and serum urea in elderly cats has been reported.6 9 11 However, in our study, no age-effect on sCr was noted.

Age-related effects on time-based parameters or parameters related to the slope of the time-intensity curve were not observed in this study. Renal resistance is reported to increase in ageing people, commonly noted as an increase in intrarenal Doppler indices.27 Nevertheless, a significant age-effect on the resistive index was also not detected in a study on 24 healthy cats.28

A significantly higher SBP was present in the cats older than 10 years compared with cats ageing between 1 and 3 years. Blood pressure in the group of 3–6 years was also higher compared with the younger cats. An age-effect on SBP has previously been described in several studies in cats.6 11 29 Efforts were made to reduce stress-related hypertension due to the ‘white coat effect’; however, single SBP measurements as done in this study are not sufficient to diagnose hypertension.19 An underlying cause for hypertension was not found by the basic work-up included in this study. Further workup and serial assessment of SBP was beyond the scope of our study. Additionally, blood pressure was measured after a short acclimatisation period in the clinic, at the beginning of the clinical examination, preceding all subsequent manipulations. It may be assumed that blood pressure may have increased by the time CEUS was performed, therefore this could have influenced the renal perfusion parameters.

The main limitation of this study is the low number of cats in each group, and even more the low number of geriatric cats.

In conclusion, no significant age-related effects on renal perfusion parameters obtained with CEUS were detected in this study. However, a tendency towards lower PE and WiAUC, potentially indicative for a lower blood volume, was present in geriatric cats. Further studies including a larger number of geriatric cats are needed to confirm these findings and determine the clinical relevance of these findings.

Acknowledgments

The authors would like to thank Bracco Suisse SA (Geneva, Switzerland) for their scientific support on the use of VueBox and Medvet (Antwerp, Belgium) for the laboratory analyses.

References

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Footnotes

  • Funding This study was funded by Bijzonder Onderzoeksfonds (2015003301).

  • Competing interests None declared.

  • Ethics approval The study protocol was approved by the local ethical committee of the Faculty of Veterinary Medicine, Ghent University, Belgium (EC2015/68) and the Deontological Committee of the Belgian Federal Agency for the Safety of the Food Chain.

  • Provenance and peer review Not commissioned; externally peer reviewed.

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