transitional cell carcinoma in the urinary bladder of a ... · m0 (tumour invading the bladder wall...

Transitional cell carcinoma in the urinary bladder of a common dolphin (Delphinus delphis) with emphasis on imaging diagnosis

Josep M. Alonso-Farré1,2, Manuel Gonzalo-Orden3, María Llarena-Reino1,4,

Tània Monreal-Pawlowsky5, Eduard Degollada6

1 Centre for Environmental and Marine Studies (CESAM),

University of Aveiro, Campus Universitário de Santiago, Aveiro, Portugal. 2Galician Stranding Network-Coordinadora para o Estudio dos Mamíferos Mariños (CEMMA)

Gondomar, Spain. 3 Department of Animal Medicine, Surgery and Anatomy, Veterinary School,

University of León, Campus de Verganzana, Spain. 4 ECOBIOMAR, Instituto de Investigaciones Marinas, IIM-CSIC, Vigo (Pontevedra), Spain.

5 Parc Zoològic de Barcelona, Barcelona, Spain 6 EDMAKTUB & Anatomy and Embryology Department, Veterinary School,

Autonomous University of Barcelona (UAB), Bellaterra, Spain.

Abstract An adult male common dolphin Delphinus delphis cadaver

was studied using ultrasound, computed tomography and

magnetic resonance imaging and subsequently frozen and

cross-sectioned for anatomical description purposes. Imaging of an

abnormal mass in the urinary bladder was seen using ultrasound and

magnetic resonance. Anatomical sections and gross dissection al-

lowed macroscopic confirmation of the mass, and histological

analysis characterized it as a transitional cell carcinoma. A purulent

abscess was also detected in the abdominal wall of the same

dolphin, by the three used imaging techniques. In cetacean species,

tumours are not reported frequently and have been considered as

rare events, but realistic cancer incidence rates are most likely

largely unknown. In this context, every case reported, as well as the

description of techniques which increase the accuracy of

diagnostics, should be considered essential.

[JMATE. 2013;6(1):11-18]

Keywords: Magnetic resonance imaging, ultrasound, computed

tomography, neoplasia, marine mammals

Introduction In marine mammals, neoplasia is not frequently

reported (10,20), with the exception of two populations

for which health status is accurately monitored: St.

Lawrence Gulf belugas Delphinapterus leucas (17) and

California sea lions Zalophus californianus (11).

Focusing on the urogenital system, carcinomas are

considered endemic in California sea lions (11), but

only two descriptions of cetacean urinary bladder

neoplasia can be found in the scientific literature, both in

belugas (15,16).

On the other hand, the diagnostic imaging

technology applied to veterinary medicine is considered

an excellent tool for neoplasia diagnosis, allowing

accurate definition of tumor margins and greater

sensitive for detecting metastasis (28). In the last years,

ultrasound (US), computed tomography (CT) and

magnetic resonance imaging (MRI) are increasingly

being used for clinical, pathological and anatomical

purposes in marine mammals (5,26). Despite the

increased use of modern technology, only one

diagnostic imaging study of a neoplasm has been

described in cetaceans to date: a poorly differentiated

brain carcinoma in a beluga by MRI (24).

Realistic neoplastic tumour incidence rates in

cetaceans could be considered largely unknown due to

the lack of epidemiological studies directly oriented to

cancer lesions in these marine species (19), except in the

case of St. Lawrence belugas. Contrary to the case of

companion animals, in which the discipline of oncology

has widely grown, in marine mammals we are in the

process of discovering the type and range of neoplastic

lesions as well as potential aetiologies. In this context,

every single case reported, as well as the description of

the techniques used for an accurate diagnosis, should be

considered essential. We present here the detection of a

carcinoma in the urinary bladder of an adult common

dolphin Delphinus delphis by means of US and MRI,

emphasizing the value of imaging techniques in marine

mammal diagnostic procedures.

Received July2, 2013; Accepted July 9, 2013

Correspondence: Josep M. Alonso-Farré

Centre for Environmental and Marine Studies (CESAM), University

of Aveiro, Campus Universitário de Santiago. 3810-193, Aveiro,

Portugal.

Phone: 0034986231930 (ext. 181)

Email: [email protected]

Journal of Marine Animals and Their Ecology Copyright © 2008 Oceanographic Environmental Research Society

Vol 6, No 1, 2013 Printed in Canada

JMATE 11

mailto:[email protected]

Material and Methods

An adult male common dolphin cadaver with a

total body length of 195 cm was fully examined by US,

CT and MRI at the Veterinary School of León (Spain).

The dolphin was found stranded on a beach in

northwestern Spain with evidence of a recent death less

than 48 hours before (13). Imaging equipment used

included an ultrasound device RT 3600 (General

Electrics Healthcare, Buckinghamshire, UK) with a 4.5

MHz probe, a single slice CT scanner Tomoscan M-EG

(Philips Medical Systems, Andover, USA), and a 0.2

Tesla Signa Profile HD (General Electrics Healthcare¸

Buckinghamshire, UK) MRI. For MRI scan acquisition,

a whole body coil was used to receive the signal, using

Fast Recovery Spin Echo sequences in T1 and T2-

weighted modes (T1W and T2W) with a field of view

(FOV) of 28, as well as pulse repetition time (TR) and

echo time (TE). Slice thickness and spacing were 4.0

mm and 1.2 mm respectively, and the image acquisition

matrix was 512x512. MRI scans in transverse, sagittal

and dorsal planes were obtained. As a general protocol,

CT scans were acquired in transverse slices of 1mm

thickness at 120 kV. Sagittal and dorsal planes could

also be examined thanks to the software image analysis.

Two algorithms (soft tissues and bony tissues) were used

during CT scanning.

After imaging, the animal was frozen at -24ºC in

ventral recumbency and then cross-sectioned with a

band saw in its transverse plane at 1 cm intervals, to

carry out a cross-sectional anatomy study (2).

Although sampling from cross-sections was possibly

bound to offer low quality samples for histological

analysis due to the freezing/thawing and cross-sectioning

processes, apparently abnormal tissues noted on images

were taken from cross-sections and preserved in 10%

buffered neutral formalin. A haematoxylin and eosin

staining protocol was used to stain histological sections

of selected tissues.

Results During the ultrasound examination of the caudal

quarter of the abdominal cavity, some abnormal

structures were identified. A noticeable, invasive, and

irregular tumour emerging from the walls of the urinary

bladder into the lumen was clearly observed. The

irregular margins of the tumour were very well defined

due to the contrast between the anechoic urine and the

surrounding soft tissues (Figures 1a and 1b). The

tumour was seen at the cranial portion of the bladder

occupying the ventral and lateral walls, affecting also

the caudal dorsal wall. The tumour occupied one third

of the cranial half and two thirds of the caudal half.

Dorsal rectal lymph nodes were enlarged compared to

normal size nodes in healthy common dolphins

(Figure 1a). An abnormal mass located in the abdominal

wall at the right side of the dolphin was also observed at

bladder level. Maximal dimensions were approximately

10x14x10 cm (left-right, dorsoventral and

craniocaudal), and it was located deep to the blubber

layer. Its ultrasonographic pattern was diffusely

hyperechoic in relation to the adjacent muscles

(Hypaxialis lumborum and Pubocaudalis), providing a

sharper identification of its margins (Figure 1c).

The bladder tumour was seen on MRI scans at the

cranial half of the bladder due to the contrast of signal

intensities obtained from urine, clearly hypointense in

T1W (Figure 2a) and hyperintense in T2W (Figure 2b),

compared to soft tissues (including the mass). Caudally,

the tumour produced a very similar signal intensity to

urine and could not be well defined, except for an

internal rounded portion of 1 cm in diameter, producing

a very hypointense signal compared to the surrounding

mass (Figure 3a). The rectal lymph nodes and rectum

could be identified in T2W MRI scans due to the lower

intensity of the signal compared with the adjacent

tissues (Figure 3a). Additionally, a craniocaudal and

dorsoventral extension of the tumour could be assessed

by means of sagittal MRI images (Figure 2c). The

lateral mass was clearly observed in T2W MRI

transverse series given that the lesion was hyperintense

in comparison to the adjacent abdominal wall muscles

(Figure 3a).

The bladder mural tumour and other

intra-abdominal organs could not be identified on CT

scans due to similar CT attenuation (Figure 3b).

However the lateral abdominal defect could be seen and

measured because of contrast in attenuation relative to

the adjacent musculature (Figure 3b).

Anatomical transverse cross-sections of the

dolphin corresponding to the urinary bladder confirmed

the existence of a pale-brown tumour of soft tissue

JMATE 12


Imaging of a common dolphin bladder carcinoma

emerging from the urinary bladder wall with a firm

(mineralized) rounded central portion 1 cm in

diameter (Figure 3c), as well as the enlargement of the

rectal lymph nodes and a purulent abscess on the lateral

abdominal wall. Several parasitic larval cysts (0.5-1 cm)

were also seen embedded in the blubber and

subcutaneously around the genital area. No other lesions

were observed in any other organ of the dolphin on

imaging or on gross dissection.

Histologically, several areas of autolysis and

disorganized tissues were observed in all the samples

studied. This was due to freezing, cross-sectioning,

thawing and collection from anatomical sections.

Despite this, bladder samples showed an enlarged

mucosa with the transitional epithelium of the smooth

muscle with more layers than normal and formed by

JMATE 13

Figure 1a: Transverse sonogram (4.5 MHz) of the caudal

abdominal cavity at the middle urinary bladder level. Probe

location: left flank (see upper dolphin icon for references).

B: blubber, HL: Hypaxialis lumborum muscle, LN: lymph nodes, R:

rectum, T: tumourous mass, U: urine (not frozen).

Figure 1b: Sagittal sonogram (4.5 MHz) of the caudal abdominal

cavity at the urinary bladder level. Probe location: ventral midline

(see upper dolphin icon). White arrows: tumourous mass,

B: blubber, RA: Rectus abdominis muscle, U: urine (not frozen).

Figure 1c: Transverse sonogram (4.5 MHz) of the caudal

abdominal cavity at the urinary bladder level. Probe location: right

flank (see upper dolphin icon). LA: Lateral abscess,

HL: Hypaxialis lumborum muscle, PC: Pubocaudalis muscle.



neoplastic cell sheets. These neoplastic cells were

characterized by dark nuclei and acidophilic cytoplasm,

and occasional abnormal mitosis was observed. No

areas of neoplastic extension were identified beyond the

muscular wall of the bladder. All of these findings sup-

port the histological diagnosis of a transitional cell

carcinoma. Tumour lesions were not seen in the lymph

nodes which presented reactive hyperplasia. The

samples of the lateral abscess only showed necrotic

debris. The tumour/node/metastasis or TNM

classification of the tumour was established as T2/N0/

M0 (Tumour invading the bladder wall with induration/

Regional lymph nodes not involved/No evidence of

distant metastasis), according to the scheme for canine

urinary bladder cancer defined by the World Health

Organization (22).

JMATE 14

Figure 2a: Transverse T1W MRI of the caudal abdominal

cavity at the cranial urinary bladder level (see upper

dolphin icon). B: blubber, BO: small intestine,

HL: Hypaxialis lumborum muscle, LV: lumbar vertebrae,

T: tumourous mass, U: urine (not frozen).

Figure 2b: Transverse T2W MRI of the caudal abdominal

cavity at the cranial urinary bladder level (see upper

dolphin icon). B: blubber, BO: small intestine,

HL: Hypaxialis lumborum muscle, LV: lumbar vertebrae,

T: tumourous mass, U: urine (not frozen).

Figure 2c: Sagittal T1W MRI of the abdominal cavity,

approximately through the midline (see upper dolphin

icon). AC: abdominal cavity, B: blubber,

RA: Rectus abdominis muscle, T: tumourous mass,

U: urine (not frozen), VC: vertebral column.



JMATE 15

Figure 3a: Transverse T2W MRI of the abdominal cavity

at the caudal urinary bladder level (see upper dolphin

icon). B: blubber, C: central portion of hard tissue,

HL: Hypaxialis lumborum muscle, LA: lateral abscess,

LN: lymph nodes, LV: lumbar vertebrae, R: rectum,

RA: Rectus abdominis muscle.

Figure 3b: Transverse CT scan of the abdominal cavity

at the caudal urinary bladder level (see upper dolphin

icon). AC: abdominal cavity, B: blubber,

HL: Hypaxialis lumborum muscle, LA: lateral abscess.

Figure 3c: Transverse anatomical cross-section of the

caudal abdominal cavity at the caudal urinary bladder

level (see upper dolphin icon). White arrows: tumourous

mass, black arrow: central portion of hard tissue,

B: blubber, HL: Hypaxialis lumborum muscle,

LA: lateral abscess, LN: lymph nodes,

LV: lumbar vertebrae, P: parasite larval cyst, R: rectum,

RA: Rectus abdominis muscle, U: urine (frozen).



Discussion

Transitional cell carcinoma of the urinary bladder

has been documented as the most common malignant

tumour of the urinary tract in other mammal species

such as the dog (18,21) or humans (25). The only case of

a transitional cell carcinoma reported in a cetacean

species occurred in one population of belugas inhabiting

the St. Lawrence estuary, Canada (15). In this area the

incidence of cancer in belugas approaches or perhaps

even exceeds that of humans (8,17), being related to the

high level of agricultural and industrial pollution (8,9).

Furthermore, urogenital carcinomas have been reported

as endemic in California sea lions, and are considered a

common cause of mortality in free-ranging adults (11).

The etiology of this neoplasia in this species is

considered likely multifactorial with several contributing

factors: viral infections (6,12), alterations in endogenous

hormone expression (7), genetic factors (3) and exposure

to pollutants (29). Although high levels of some

pollutants have been described in tissues from common

dolphins of NW Spain (23), the case presented here is

the first diagnosed cetacean neoplasia in this

geographical area.

Abscesses embedded in the blubber and

subcutaneous tissue around the genital area are

frequently observed in dolphins stranded in NW Spain,

most of the times associated with cestode

Phyllobothrium delphini cysts degeneration (1). These

processes commonly form smaller lesions than those

observed in our case, but in some cases they may cause

large purulent abscesses, especially in deeply weakened

and immunosupressed dolphins. On the other hand,

metastases of urogenital carcinomas in California sea

lions are found throughout pelvic and abdominal lymph

nodes, as well as abdominal and thoracic viscera (7).

These metastatic lesions could also degenerate

producing necrotic lesions and abscesses. In our case,

definitive characterization of the lateral abscess origin

was not possible due to the insufficient quality of

samples obtained after the anatomical procedures.

Diagnostic imaging techniques such as US, CT

and MRI applied to animal medicine are considered

excellent tools for the diagnosis of neoplasia, allowing

an accurate definition of tumour margins and effective

detection of metastasis (28). The US and MRI images

obtained in our case clearly revealed the invasive

tumour, occupying half of lumen of the urinary bladder.

Even though the definition of the lesions nature is not

permitted by any imaging technique, the imaging

patterns of the tumour presented here are coincident

with those of TCC in dogs (14) or humans (27). Both

techniques allowed transverse and sagittal imaging

planes which were found to be essential for an accurate

assessment of the tumour extension.

As in terrestrial mammals, urinary bladder

proliferative lesions in cetacean species can be

asymptomatic or provoke obstructive and/or irritative

symptoms, hematuria, and flank pain. As these

symptoms are considered non-specific, imaging

techniques are valuable diagnostic tools to achieve more

accurate diagnostics. Traditional diagnostic imaging of

the urinary bladder included contrast cystography or

intravenous urography (IVU). As described here, US

and MRI could be alternative or complementary meth-

odologies, with the additional value that they do not

produce ionizing radiation exposure. Furthermore, CT-

IVU has been successfully used to visualize the urinary

bladder, but tissue could not be reliably distinguished

from fluid without IV contrast which is not possible in

post-mortem CT scans. US has been described as an

ideal imaging tool for male dolphin’s genital-urinary

tract examination, allowing a clear image of the urinary

bladder (4). It is a non-ionizing, low cost, fast and easily

repeatable imaging technique in which no

tranquilization or general anaesthesia of the patient is

required. In the last two decades, US portable devices

have been developed and nowadays, they can be easily

transported and used very close to marine mammal

pools, permitting comfortable handling and reducing the

stress level of the cetacean. The use of color Doppler to

distinguish mural masses from avascular luminal

sediment or hematomas is valid for living animals but

not an option when dealing with cadavers. On the other

hand, the use of CT and MRI in cetaceans is currently

still limited because scanners are often not available and

transport of cetaceans to the location of such equipment

is not without risks. Nevertheless, the interest in moving

forward in its use is considerable, especially when

dealing with endangered species.

Finally, the use of imaging techniques in dead

dolphins considerably improves post-mortem

information acquisition and allows us to gather

JMATE 16



pathological, anatomical and morphological data from

museum specimens or dolphins that cannot be fully

necropsied. This may be a key factor for increasing the

low number of reported tumours and for widening the

current knowledge of the real incidence of neoplasia in

cetacean species.

Acknowledgements

The authors want to thank stranding networks staff of Galicia

(CEMMA) for technical support to this research. We want also

thank Fiona Read for her constructive comments on the manuscript.

First author is currently funded under post-doctoral fellowship

SFRH/BPD/47251/2008 of the Fundação para a Ciência e a

Tecnologia (Portugal).

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