J. Vet. Med. A 39, 759-768 (1992) 0 1992 Paul Parey Scientific Publishers, Berlin and Hamburg ISSN 0931 - 184X
From the Center of Biomedical Research and the II. Surgical Department, University of Vienna, and from the Institute of Anatomy, Animal Genetics and II. Department of Internal Medicine, Veterinary University of Vienna, Austria
Cardiac Function, Morphology and Chromosomal Aberrations in a Calf with Ectopia Cordis Cervicalis URSULA WINDBERGER, G. FORSTENPOINTNER, F. GRABENWOGER, E. KOPP,W. KUNZEL, B. MAYR,A. PERNTHANER, P. SIMONand U. LOSERT Address of authors: Dr. URSULA WINDBERGER, Center for Biomedical Research, University of Vienna, General Hospital, Wahringergiirtel 18-20, A-1090 Vienna
With 5 figu~es ~ OctobeY 3, 1992) (Received f o publication
Summary A male calf with ectopia cordis cervicalis inferior was investigated clinically, hemodynamically, and morphologically from the 3rd day of age until slaughter at the age of 22 months. Arterial hypertension (mean AOP 140 mm Hg), concentric myocardial hypertrophy and good ventricular contractility with normal valve function were found. Normal clinical condition at rest (cardiac output 48 Vmin in the 600 kg bullock), but cyanosis and dyspnoe during physical exercise were observed. Variations in the large vessels, such as persistence of the right cranial caval vein and one common pulmonary vein emptying into the left atrium, were found. Abrupt caliber differences of large arterial vessels together with a right angled aortic arch were observed, presumably contributing to increased afterload. The cytogenetic analysis showed the presence of a small marker chromosome leading to a chromosome number of 61 in a fraction of metaphases (mosaicism). Moreover, chromosome breakages were observed in many cells. From the 5 cows which were inseminated, 3 became pregnant. Ectopia cordis was not observed in any offspring. We conclude that in this animal compensated cardiac insufficiency with peripheral hypertension was established.
Introduction Ectopia cordis is defined as an abnormal position of the heart outside the thoracic cavity. It can be divided into the superior and inferior cervical, the sternal, and the abdominal types. Since information on the cardiac function in ectopia cordis cervicalis is limited, we describe the clinical and hemodynamic parameters together with morphological abnormalities in a male calf, breed “Fleckvieh”. Additionally, we tested for chromosomal aberrations and report the clinical outcome of 3 offspring.
Methods Clinical Observation and Hernodynamic Tests During the calf‘s life span until slaughtering at the age of 21 months, it was weighed and clinically examined (13) at weekly intervals. U.S.Copyright Clearance Center Code Statement:
0931 - 184X/92/3910-0759$02.50/0
760
URSULA WINDBERGER et al.
At the age of 3 weeks, under general anaesthesia (thiopentone/atropin; halothane / nitrous oxide), a catheter was inserted into the left ventricle via the left carotid artery. A second catheter had to be placed in the left cranial caval vein because it was not possible to insert it into the right heart. Both catheters were connected to a Statham pressure transducer for on-line monitoring of the left ventricular pressure (LVP), aortic pressure (AOP) after retraction of the catheter from the left ventricle into the aorta, and central venous pressure (CVP). The transducer was adjusted at the height of the mitral valve although difficulties existed in the identification of this level because of significant cardiac motion. Additionally, extremity leads for E C G were attached in the standard way. Venous blood was withdrawn and common hematologic, blood coagulation, blood chemical parameters, and an electrophoresis of plasma proteins (Hitachi 737 and 705, Tokyo, Japan; Sysmex CC 800, Toa Medical Electronics Ltd., Kobe, Japan) were measured. For the conventional angiographic recording, Jopamiro 300 was injected into the jugular vein and in a second series into the left ventricle. Three days following this intervention, the conscious calf was echocardiographically examined (Ultramark 4, Advanced Technical Laboratories, Washington District, USA) in upright position. As soon as the calf was calm for 30 minutes, the arterial pressure was monitored on-line in the ear artery and a blood gas analyzed (blood gas analyzer 1306, Intrumentation Laboratory, Milano, Italy). A further X-ray examination of the conscious and standing calf was performed at the age of 6 weeks. At the age of 20 months the animal was sedated with 60 mg xylazine and a second echocardiogram (Sonos 1000, Hewlett-Packard, Andover, USA) performed. The arterial pressure was monitored from the ear artery as before.
Morphologic Investigations The bullock was sacrificed at the age of 21 months. The anatomy of the major vessels at the base of the heart were determined during dissection. The heart and lung were investigated pathohistologically and biopsies of the M m . soleus, tibialis cranialis and extensor digitorum longus were investigated histochemically for muscle morphology after staining with ATPase ( p H 10.4 and 4.5) and N A D H (3, 4). Cytogenetic Analysis G-banding was done according to WANGand FEDEROFF (24). Semen was withdrawn from the Ductus epididyrnidis and 5 cows were inseminated. The offspring were examined clinically at the age of 7 days.
Results Clinical and ffemodynamic Results During the clinical investigation at the age of 3 days, the pulse was strong and regular at a rate of 80 b/min. The calf suffered from E . coli enteritis, which responded well to antibiotic therapy. The X-ray examination showed increased vascularity in the lungs and kinking of the munubriurn sterni. The anatomic heart axis pointed from caudodorsal to cranioventral. In the anaesthetized animal the LVP (190 mm Hg) was elevated and irregular. Due to the exaggerated motion of the heart inside loose connective tissue there was a swing in the LVP. The AOP was increased (150mm H g mean) and the pulse pressure of the A O P differed from one beat to the other but was about 20-30mm Hg. The heart rate also appeared irregular. CVP mean was 13 mm H G (Fig. 1). The mean arterial pressure monitored from the ear artery in the conscious calf after 3 days was 145mm Hg. The heart rate was 140b/min. The myocardial contractility was estimated semi-quantitatively by the echocardiographic investigation and was found to be normal with normal valve function. Peak aortic flow velocity was 2 m/sec. Normal values were received from all laboratory examinations (15). From birth through 20 months the animal showed a daily increase in live mass of 1 . 1 kg. In the last month the body weight decreased by 30 kg and when sacrificed the bullock weighed 640 kg. During that time, variable symptoms of bronchitis were observed which did not completely respond to antibiotic and antiinflammatory therapy.
Cardiac Function, Morphology and Chromosomal Aberrations in a Calf
76 1
Fig. 1. ECG, LV dp/dt, left ventricular pressure (LVP), aortic pressure (AOP), and central venous pressure (CVP) during general anaesthesia
In the resting calf, a normal clinical condition with a strong pulse at a rate of 80 b/min was seen which increased during physical exercise up to 140 b/min, and dyspnoe and cyanosis were observed. Auscultation of the extrathoracic heart revealed normal findings. The mean arterial pressure measured in the ear artery was 150mm Hg. The echocardiographic examination at the age of 20 months showed normal contractility without valvular regurgitation. Left ventricular end diastolic diameter was 92 mrn with an end systolic diameter of 54mm. The shortening fraction of the left ventricle was 41 % and thickening fraction of the interventricular septum was 40 % (thickness of the septum in diastole: 18mm, in systole: 30mm). The diameter of the sinus valsalvae was 73mm and that of the aorta ascendens 52 mm. The cardiac output was calculated by multiplying the echocardiographically determined stroke volume (875 ml) by the heart rate (55 blrnin) and it was 48 Vmin. Peak doppler aortic flow velocity was 170cm/sec with a mean velocity of 118 crn/sec. Molphological Results The heart was located level with the 5th-7th cervical vertebra ventrally to the trachea. The apex cordis was directed cranioventrally, the facies auricularis was on the left, the facies atrialis on the right side. The heart was completely wrapped in the pericardiurn which was strongly adherent to the surrounding tissues. Cardiac muscle cells were seen in the terminal portion of the 6 cm V. cava cranialis dextra. The V. azygos dextra flowed dorsally into the V. cava nun. dext. directly before its opening into the right atrium. 3.5 cm cranial thereafter splitting into the V. subclavia dextra and the V. jugularis externa dextra. Caudally to the intervenous tubercel was the opening of the V. cava cuudalis and ventrally the opening of the Sinus coronarius of approximately the same size. The Sinus coronarius ran caudally from the left atrium to the left side and continued dorsocranially forming the V. cava cranialis sinistra. Into this vessel cranially the thin V. jugularis externa sinistra, dorsocaudally the V. azygos sinistra and craniodorsally the V. costocervicalis sinistra were
762
URSULAWINDBERGER et al.
joined. The pulmonary trunk, connected to the aortic arch by the persistent arterial duct, produced the longitudinally extended Aa. pulmonalis dextra and sinistra (Fig. 2). Only one wide-lumened V.pulmonalis opened into the left atrium. The interior spaces of the atrium sinistrum and the auricula sinistra showed no abnormalities. The aorta ran straight dorsally forming a nearly right angled aortic arch next to the trachea. A 10 cm Truncus bracbiocepbalicus extended from the aortic arch. This TY.bracbiocepbalicus was enlarged and formed first the A . carotis communis sinistra and then, originating from one common trunk, both the A. carotis communis dextra and the A. subclavia dextra, which
Fig.2. Angiogram of the right heart with the venous inflow and the pulnomary artery. 1 V .jugularis extena sinistra; 2 - V.subclavia sinistra; 3 - V . azygos sinistra; 4 - V . costocenticalis sinistra; 5 - V . cava cranialis sinistra; 6 - Atrium dextrum; 7 - Auricuh dextra; 8 - Ventriculus dexter; 9 - Truncus pulmonalis; 10 - Catheter in A . carotis communis sinistra
Cardiac Function, Morphology and Chromosomal Aberrations in a Calf
763
descended from the Truncus vertebrocostocervicalis dexter. At the end of the aortic arch the
A. subclavia sinistra, and from its onset a Truncus vertebrocostocervicalis sinister branched off (Fig. 3). Both ventricles demonstrated myocardial hypertrophy. Fiber degeneration and myocarditis simplex were seen both in the ventricles, and mainly in the atrial myocardium. No pecularities were observed in the interior spaces of the right or left chambers of the heart. In patches throughout the lung, emphysema and atelectasis combined with follicular peribronchitis was observed. In the M . tibialis cranialis 25 % oxidative type I and 75 %
Fig. 3. Angiogram'of the left heart and the great arterial vessels. 1 - Catheter in A . carotis communis sinistra; 2 - Ventriculus sinister; 3 - Ventricubs sinister; 4 - Bulbus aortae; 5 - Aorta descendens; 6 - Truncus brachiocephalicus; 7 - A.carotis communis dextra; 8 - A.subclavia dextra; 9 - A . subclavia sinistra; 10 - Truncus vertebrocostocervicalis dexter; 11 - Truncus vertebrocostocervicalis sinister; 12 - Catheter in V .jugularis externa sinistra
764
URSULA WINDBERGER et al.
Fig.4. G-banded (a-f) metaphases of the investigated ectopic bull. a-d: metaphases with a chrornosome number of 61. Note the small marker chromosome (M); e-f: G-banded metaphases showing the breakages (arrows). Bars represent 10 pm
Cardiac Function, Morphology and Chromosomal Aberrations in a Calf
765
glycolytic type I1 b fibers were found in sections stained for actomyosin ATPase and N A D H indicating a quick contracting muscle with easy fatigue-ability. In contrast, the M . extensor digitorum longus showed 90 % oxidative fibers with about 50 % quick (type
Fig. 5. Katyotype of metaphase of Fig. 4 b. Note the marker chromosome (M). Bar represents 10 pm
766
URSULA WINDBERGER et al.
I1 a) and 50 % slow contracting (type I) fibers. The M . soleus contained 60 % oxidative type I fibers and about 40 % oxidative type I1 a or I1 x fibers. Both muscles were identified as mainly slow and fatigue resistant.
Cytogenetic Results In 20.5 % of the examined metaphases (41 of 200 cells), cytogenetic alterations were observed. In about a half of these altered cells (18 cells) a small marker chromosome was observed (Fig. 4 a, b, c, d and Fig. 5 ) in addition to the bovine diploid chromosome set (2 n = 60). Thus, 61 chromosomes were present in these cells. Moreover, chromosome breakages and fragments predominated among the cytogenetic alterations (Fig. 4 e and f). No such alterations were oberserved in simultaneously performed control cultures. From the 5 cows which had been inseminated, 3 became pregnant. Ectopia cordis was not observed in any of the newborns. Two calves had a normal clinical picture. One calf died 1 day post partum from unknown causes. Discussion Reports of ectopia cordis in man originate in 1671 (22), however, the first case in veterinary literature was published in 1818 (2). Since then, numerous descriptions have been made concerning morphologic abnormalities in calves with ectopia cordis. Whereas the sternal and abdominal types are found in man (24, dog ( 5 ) , pig (6), and pigeon (19), in cattle the lower cervical type seems to be most common (2, 8, 11, 12, 14, IS), although the sternal type has already been described (17). The percentage of total congenital defects which are cardiac in cattle, varies from 1 . 5 % to 9.6% and depends on breed and geographic region. For example, 1.5 '/o to 2.7 % has been reported in Germany (16, 23), whereas 1.9 YO in Kansas, USA (9), 8.3 Yo in Japan (lo), and 9.6 % in Switzerland (17) have been described. According to different authors, ectopia cordis accounts for 1 % to 45 YO of all cardiac defects in cattle: 1 % (17), 13 % (S), 15 O/O (16) and 45 % (9). Surgical correction has been tried in man (22) and cattle (I), but has been entirely unsuccessful. We describe a calf with ectopia cordis cervicalis inferior with certain anatomic abnormalities of the great vessels which result in several hemodynamic alterations. To our knowledge this is the first description of functional properties of a congenital extrathoracic heart. The position of the great vessels is comparable to that of a bovine fetus 35 days after conception (7). At that time, the embryonic heart is near the 5.-6. cervical segment and the aortic arch near the 6. segment. Due to the lack of fusion of both cranial caval veins, the V. cava man. sin. remained. The V.jugularis externa, subclavia, and azygos drain into the Vv. cavae cran. of the corresponding side of the body. The persistent aortic arch at the 6. cervical segment prevents the junction of the Tr. brachiocephalicus with the A . subclavid sin. Similar morphologic observations such as persistence of the left cranial caval vein, emptying of one common pulmonary vein into an elongated left atrium, changes in the course of the arteries, and the sharp ventral bend of the sternum have been reported previously (2, 11, 12, 18). In our study we observed massive concentric myocardial hypertrophy. The cause of the left ventricular hypertrophy is unclear and may be multifactorial. It seems possible that the abnormal course of three thin arteries, both carotid arteries and one subclavian artery, arising from an enlarged Tr. brachiocephalicus, and the rightangled aortic arch result in an elevated afterload. Furthermore, the arterial hypertension increases afterload. Both the left ventricular pressure and the arterial pressure were elevated, although information on the exact value can not be provided due to the exaggerated movement of the heart within loose connective tissue. The dp/dt of the LVP was estimated in some circumstances as being up to 8000 torr/ sec whereas most of the time normal values were measured. The irregular cardiac rhythm together with the elevated heart rate and the increased LVP would contribute to this increased dp/dt during several cardiac cycles. Following the retraction of the catheter from
Cardiac Function, Morphology and Chromosomal Aberrations in a Calf
767
the left ventricle into the aorta a gradient of about 40 mm Hg was observed. Based on the observation that both the length of the aorta and its diameter, at least in the Sinus valsaluae and after the right angled aortic arch, was increased in this animal (see Fig. 2), an increase of the aortic compliance may be presumed. This increased compliance may also explain the low pulsatility of the aortic pressure. Furthermore, functional aortic stenosis following luxation of the heart, swinging during the cardiac cycles, might also contribute to the observed effect. Nevertheless, the LVP as well as the peripheral arterial pressure were elevated. The etiology of the hypertension is unclear. It may be an essential hypertension or may be due to altered baroreceptor sensitivity secondary to the abnormal great vessel anatomy, such as it can be seen in aortic coarctation. The hypertrophy of the right ventricle may indicate increased pulmonary vasculary resistance. Unfortunately, direct hemodynamic measurements in the pulmonary circulation could not be performed due to the kinking of the left cranial caval vein. However, since both ventricles showed myocarditis simplex, a primary muscle disease cannot be excluded. We observed that the resting calf appeared clinically normal, with normal pulse and color of the mucous membranes, whereas in a state of increased metabolic demand, even trotting, the animal showed signs of cardiac decompensation such as increase of the heart rate, cyanosis and dyspnoe. However, the function of liver and kidney was maintained. Cytogenetically, a major fraction of the investigated cells (mosaicism) of the calf revealed the presence of an additional small marker chromosome, giving rise to a chromosome number of 61. This marker possesses a clearly recognizable centromer. Moreover, the presence of two G-bands was typical for this small marker. However, a more definite cytogenetic characterisation, including an unequivocal identification, was not possible. Of further interest were the breakages observed in many metaphases. Chromosome breakage syndromes with background values of 1.8 % and 2 %, found in cattle (20, own unpublished data), play some role in several hereditary diseases at least in man. In our extopia calf, we observed a value of 20.5% of alterations which is clearly increased. Different chromosomes, in most cases hardly to be identified, were affected. High percentages of alterations (12 %, 15 Yo) have also been observed in two cases of bovine congenital harilessness (21). Similar to our present case, the increased alteration frequency could not be traced back to a certain chromosome segment. The 3 clinically normal offspring of our bullock showed a normal thoracic radiogram, in agreement with a previous report on normal offspring produced from a cow with ectopia cordis cervicalis (2). We conclude that in this case of ectopia cordis cervicalis inferior, the changed anatomic situation in the great vessels possibly led to increased afterload resulting in ventricular hypertrophy and compensated cardiac failure. Peripheral hypertension, either essential or as a result of altered baroreceptor activity, was also observed. Except for the markedly altered anatomic situation, the cardiac output was maintained at rest. Acknowledgement This study was supported by the Ludwig-Boltzmann Foundation. References 1. ASLANBEY, D., M.K. GAVUZOGLU, T.T~ssas,S.BERKIN, and B.YILMAZ, 1980: Bir danada ectopia cordis cervicalis olgusu. Veteriner Facultesi Dergisi Ankara Universitesi 27, 613 -632. 2. BOWEN, J.M., and R. W. ADRIAN, 1962: Ectopia cordis in cattle. J. Am. Vet. Med. Assoc. 141, 1162- 1167. 3. BROOKE, M. H., and K. K. KAISER,1970: Muscle fibre types: how many and what kind? Arch. Neurol. 23, 369-379. 4. BRUMBACK, R. A., and R. W. LEECH,1984: Color atlas of muscle histochemistry. Littleton, Massachusetts: PSG Publishing Company Inc., pg. 16.
768
URSULAWINDBERGER et al.
5. FREEMAN, L. E., and P. T. MCGOVERN, 1984: Thoracoabdominal ectopia cordis in two littermate pups. Canine Practice 11, 39-43. L. E., and P.T. MCGOVERN, 1984: Ectopia cordis thoracoabdominalis in a piglet. Vet. 6. FREEMAN, Rec. 115, 431-433. 7. GOLUB,D. M., 1931: Die Entwicklung des Aortenbogens und der von ihm ausgehenden Aste bei Saugetieren. I. Entwicklung der Kiemenbogen bei Kalb und Menschen. Zschr. Anat. Entw. Gesch. 95, 447-472. 8. GOPAL,T., H. W. LEIPOLD,and S. M. DENNIS, 1986: Congenital cardiac defects in calves. Am. J. Vet. Res. 47, 1120-1121. J. L. NOORDSY, and S. M. DENNIS,1973: Congenital 9. GREENE,H. J., H . W. LEIPOLD,K. HUSTON, defects in cattle. Irish Vet. J. 27, 37-45. 10. HAGIO,M., 1985: Congenital heart disease in cattle. Bull. Fac. Agric. Miyazaki Univ. 32, 233-250. 11. HIRAGA, T., and M. ABE, 1986: Eight calves of cervical ectopia cordis and their sternums. Jpn. J. Vet. Sci. 48, 1199- 1206. 12. HUGHES,H.V., 1934: Ectopia cordis in the calf. J. Comp. Path. Ther. 47, 141-151. 13. JAKSCH,W., und E. GLAWISCHNIG, 1981: Klinische Propadeutik der Inneren Krankheiten und Hautkrankheiten der Haustiere. Paul Parey Scientific Publishers, Berlin and Hamburg, 2nd edition. 14. JANIAK, T., 1954: Wrodzone przemieszczenie serca na szyje u rocznego buhaja. Med. Wtryn. 10, 83 - 86. 15. KANEKO, J. J. (ed.), 1980: Clinical biochemistry of domestic animals. Academic Press, New York - London - Toronto - Sydney - SanFrancisco, 3rd edition. H. H., 1985: Statistische Analyse des Auftretens von Kalbern mit kongenitalen 16. KOMOROWSKY, Anomalien in der Nachkommenschaft von Besamungsbullen in Hessen, eine explorative Datenbeschreibung. Dissertation Justus-Liebig-Univ. GieBen. und R. FATZER,1980: Angeborene morphologische Anomalien bei Kalbern 17. KONIG,B., A. TONTIS aus dem Raum Bern. Schweiz. Arch. Tierheilk. 122, 435-458. 18. KULCZYCKI, W., 1902: Ein Fall von Ectopia cordis beim Kalbe. Polnisches Archiv fur biologische und medizinische Wissenschaften 1, 1 - 15. and K. MEYER,1983: Ectopia cordis bei einer Taube. Dtsch. 19. KUMMERFELD, N., U. NEUMANN, tierarztl. Wschr. 90, 179- 180. 20. LOJDA, L., J. RUBES,and J. MRAZ, 1984: Levels of spontaneous chromosome aberrations in populations of cattle and pigs in Czechoslovakia as indicators of environmental mutagenic effects. Proceedings of the 6th European Colloquium o n Cytogenetics of Domestic Animals, Zurich, 104-106. 21. MAYR, B., 1979: Untersuchungen zytogenetischer Marker und ihrer Anwendbarkeit in der Zuchtpraxis bei Rind und Schwein. Habilitationsschrift, Veterinarmed. Universitat Wien. 22. MILLHOUSE, R. F., and H.A. JOOS, 1959: Extrathoracic ectopia cordis: report of cases and review of literature. Am. heart J. 57, 470-476. 1972: Untersuchungen zur teratologischen 23. RIECK, G. W., K. H. FINGER und A. HERZOG, Populationsstatistik und 7ur Atiologie der embryonakn Entwicklungsstorungen beim Rind. 1. Teratologische Populationsstatistik. GieDener Beitr. Erbpath. Zuchthyg. 4, 39-69. 24. WANG,H.C., and S.FEDOROFF,1972: Banding in human chromosomes treated with trypsin. Nature New Biol. 235, 52-54.
From the Center of Biomedical Research and the II. Surgical Department, University of Vienna, and from the Institute of Anatomy, Animal Genetics and II. Department of Internal Medicine, Veterinary University of Vienna, Austria
Cardiac Function, Morphology and Chromosomal Aberrations in a Calf with Ectopia Cordis Cervicalis URSULA WINDBERGER, G. FORSTENPOINTNER, F. GRABENWOGER, E. KOPP,W. KUNZEL, B. MAYR,A. PERNTHANER, P. SIMONand U. LOSERT Address of authors: Dr. URSULA WINDBERGER, Center for Biomedical Research, University of Vienna, General Hospital, Wahringergiirtel 18-20, A-1090 Vienna
With 5 figu~es ~ OctobeY 3, 1992) (Received f o publication
Summary A male calf with ectopia cordis cervicalis inferior was investigated clinically, hemodynamically, and morphologically from the 3rd day of age until slaughter at the age of 22 months. Arterial hypertension (mean AOP 140 mm Hg), concentric myocardial hypertrophy and good ventricular contractility with normal valve function were found. Normal clinical condition at rest (cardiac output 48 Vmin in the 600 kg bullock), but cyanosis and dyspnoe during physical exercise were observed. Variations in the large vessels, such as persistence of the right cranial caval vein and one common pulmonary vein emptying into the left atrium, were found. Abrupt caliber differences of large arterial vessels together with a right angled aortic arch were observed, presumably contributing to increased afterload. The cytogenetic analysis showed the presence of a small marker chromosome leading to a chromosome number of 61 in a fraction of metaphases (mosaicism). Moreover, chromosome breakages were observed in many cells. From the 5 cows which were inseminated, 3 became pregnant. Ectopia cordis was not observed in any offspring. We conclude that in this animal compensated cardiac insufficiency with peripheral hypertension was established.
Introduction Ectopia cordis is defined as an abnormal position of the heart outside the thoracic cavity. It can be divided into the superior and inferior cervical, the sternal, and the abdominal types. Since information on the cardiac function in ectopia cordis cervicalis is limited, we describe the clinical and hemodynamic parameters together with morphological abnormalities in a male calf, breed “Fleckvieh”. Additionally, we tested for chromosomal aberrations and report the clinical outcome of 3 offspring.
Methods Clinical Observation and Hernodynamic Tests During the calf‘s life span until slaughtering at the age of 21 months, it was weighed and clinically examined (13) at weekly intervals. U.S.Copyright Clearance Center Code Statement:
0931 - 184X/92/3910-0759$02.50/0
760
URSULA WINDBERGER et al.
At the age of 3 weeks, under general anaesthesia (thiopentone/atropin; halothane / nitrous oxide), a catheter was inserted into the left ventricle via the left carotid artery. A second catheter had to be placed in the left cranial caval vein because it was not possible to insert it into the right heart. Both catheters were connected to a Statham pressure transducer for on-line monitoring of the left ventricular pressure (LVP), aortic pressure (AOP) after retraction of the catheter from the left ventricle into the aorta, and central venous pressure (CVP). The transducer was adjusted at the height of the mitral valve although difficulties existed in the identification of this level because of significant cardiac motion. Additionally, extremity leads for E C G were attached in the standard way. Venous blood was withdrawn and common hematologic, blood coagulation, blood chemical parameters, and an electrophoresis of plasma proteins (Hitachi 737 and 705, Tokyo, Japan; Sysmex CC 800, Toa Medical Electronics Ltd., Kobe, Japan) were measured. For the conventional angiographic recording, Jopamiro 300 was injected into the jugular vein and in a second series into the left ventricle. Three days following this intervention, the conscious calf was echocardiographically examined (Ultramark 4, Advanced Technical Laboratories, Washington District, USA) in upright position. As soon as the calf was calm for 30 minutes, the arterial pressure was monitored on-line in the ear artery and a blood gas analyzed (blood gas analyzer 1306, Intrumentation Laboratory, Milano, Italy). A further X-ray examination of the conscious and standing calf was performed at the age of 6 weeks. At the age of 20 months the animal was sedated with 60 mg xylazine and a second echocardiogram (Sonos 1000, Hewlett-Packard, Andover, USA) performed. The arterial pressure was monitored from the ear artery as before.
Morphologic Investigations The bullock was sacrificed at the age of 21 months. The anatomy of the major vessels at the base of the heart were determined during dissection. The heart and lung were investigated pathohistologically and biopsies of the M m . soleus, tibialis cranialis and extensor digitorum longus were investigated histochemically for muscle morphology after staining with ATPase ( p H 10.4 and 4.5) and N A D H (3, 4). Cytogenetic Analysis G-banding was done according to WANGand FEDEROFF (24). Semen was withdrawn from the Ductus epididyrnidis and 5 cows were inseminated. The offspring were examined clinically at the age of 7 days.
Results Clinical and ffemodynamic Results During the clinical investigation at the age of 3 days, the pulse was strong and regular at a rate of 80 b/min. The calf suffered from E . coli enteritis, which responded well to antibiotic therapy. The X-ray examination showed increased vascularity in the lungs and kinking of the munubriurn sterni. The anatomic heart axis pointed from caudodorsal to cranioventral. In the anaesthetized animal the LVP (190 mm Hg) was elevated and irregular. Due to the exaggerated motion of the heart inside loose connective tissue there was a swing in the LVP. The AOP was increased (150mm H g mean) and the pulse pressure of the A O P differed from one beat to the other but was about 20-30mm Hg. The heart rate also appeared irregular. CVP mean was 13 mm H G (Fig. 1). The mean arterial pressure monitored from the ear artery in the conscious calf after 3 days was 145mm Hg. The heart rate was 140b/min. The myocardial contractility was estimated semi-quantitatively by the echocardiographic investigation and was found to be normal with normal valve function. Peak aortic flow velocity was 2 m/sec. Normal values were received from all laboratory examinations (15). From birth through 20 months the animal showed a daily increase in live mass of 1 . 1 kg. In the last month the body weight decreased by 30 kg and when sacrificed the bullock weighed 640 kg. During that time, variable symptoms of bronchitis were observed which did not completely respond to antibiotic and antiinflammatory therapy.
Cardiac Function, Morphology and Chromosomal Aberrations in a Calf
76 1
Fig. 1. ECG, LV dp/dt, left ventricular pressure (LVP), aortic pressure (AOP), and central venous pressure (CVP) during general anaesthesia
In the resting calf, a normal clinical condition with a strong pulse at a rate of 80 b/min was seen which increased during physical exercise up to 140 b/min, and dyspnoe and cyanosis were observed. Auscultation of the extrathoracic heart revealed normal findings. The mean arterial pressure measured in the ear artery was 150mm Hg. The echocardiographic examination at the age of 20 months showed normal contractility without valvular regurgitation. Left ventricular end diastolic diameter was 92 mrn with an end systolic diameter of 54mm. The shortening fraction of the left ventricle was 41 % and thickening fraction of the interventricular septum was 40 % (thickness of the septum in diastole: 18mm, in systole: 30mm). The diameter of the sinus valsalvae was 73mm and that of the aorta ascendens 52 mm. The cardiac output was calculated by multiplying the echocardiographically determined stroke volume (875 ml) by the heart rate (55 blrnin) and it was 48 Vmin. Peak doppler aortic flow velocity was 170cm/sec with a mean velocity of 118 crn/sec. Molphological Results The heart was located level with the 5th-7th cervical vertebra ventrally to the trachea. The apex cordis was directed cranioventrally, the facies auricularis was on the left, the facies atrialis on the right side. The heart was completely wrapped in the pericardiurn which was strongly adherent to the surrounding tissues. Cardiac muscle cells were seen in the terminal portion of the 6 cm V. cava cranialis dextra. The V. azygos dextra flowed dorsally into the V. cava nun. dext. directly before its opening into the right atrium. 3.5 cm cranial thereafter splitting into the V. subclavia dextra and the V. jugularis externa dextra. Caudally to the intervenous tubercel was the opening of the V. cava cuudalis and ventrally the opening of the Sinus coronarius of approximately the same size. The Sinus coronarius ran caudally from the left atrium to the left side and continued dorsocranially forming the V. cava cranialis sinistra. Into this vessel cranially the thin V. jugularis externa sinistra, dorsocaudally the V. azygos sinistra and craniodorsally the V. costocervicalis sinistra were
762
URSULAWINDBERGER et al.
joined. The pulmonary trunk, connected to the aortic arch by the persistent arterial duct, produced the longitudinally extended Aa. pulmonalis dextra and sinistra (Fig. 2). Only one wide-lumened V.pulmonalis opened into the left atrium. The interior spaces of the atrium sinistrum and the auricula sinistra showed no abnormalities. The aorta ran straight dorsally forming a nearly right angled aortic arch next to the trachea. A 10 cm Truncus bracbiocepbalicus extended from the aortic arch. This TY.bracbiocepbalicus was enlarged and formed first the A . carotis communis sinistra and then, originating from one common trunk, both the A. carotis communis dextra and the A. subclavia dextra, which
Fig.2. Angiogram of the right heart with the venous inflow and the pulnomary artery. 1 V .jugularis extena sinistra; 2 - V.subclavia sinistra; 3 - V . azygos sinistra; 4 - V . costocenticalis sinistra; 5 - V . cava cranialis sinistra; 6 - Atrium dextrum; 7 - Auricuh dextra; 8 - Ventriculus dexter; 9 - Truncus pulmonalis; 10 - Catheter in A . carotis communis sinistra
Cardiac Function, Morphology and Chromosomal Aberrations in a Calf
763
descended from the Truncus vertebrocostocervicalis dexter. At the end of the aortic arch the
A. subclavia sinistra, and from its onset a Truncus vertebrocostocervicalis sinister branched off (Fig. 3). Both ventricles demonstrated myocardial hypertrophy. Fiber degeneration and myocarditis simplex were seen both in the ventricles, and mainly in the atrial myocardium. No pecularities were observed in the interior spaces of the right or left chambers of the heart. In patches throughout the lung, emphysema and atelectasis combined with follicular peribronchitis was observed. In the M . tibialis cranialis 25 % oxidative type I and 75 %
Fig. 3. Angiogram'of the left heart and the great arterial vessels. 1 - Catheter in A . carotis communis sinistra; 2 - Ventriculus sinister; 3 - Ventricubs sinister; 4 - Bulbus aortae; 5 - Aorta descendens; 6 - Truncus brachiocephalicus; 7 - A.carotis communis dextra; 8 - A.subclavia dextra; 9 - A . subclavia sinistra; 10 - Truncus vertebrocostocervicalis dexter; 11 - Truncus vertebrocostocervicalis sinister; 12 - Catheter in V .jugularis externa sinistra
764
URSULA WINDBERGER et al.
Fig.4. G-banded (a-f) metaphases of the investigated ectopic bull. a-d: metaphases with a chrornosome number of 61. Note the small marker chromosome (M); e-f: G-banded metaphases showing the breakages (arrows). Bars represent 10 pm
Cardiac Function, Morphology and Chromosomal Aberrations in a Calf
765
glycolytic type I1 b fibers were found in sections stained for actomyosin ATPase and N A D H indicating a quick contracting muscle with easy fatigue-ability. In contrast, the M . extensor digitorum longus showed 90 % oxidative fibers with about 50 % quick (type
Fig. 5. Katyotype of metaphase of Fig. 4 b. Note the marker chromosome (M). Bar represents 10 pm
766
URSULA WINDBERGER et al.
I1 a) and 50 % slow contracting (type I) fibers. The M . soleus contained 60 % oxidative type I fibers and about 40 % oxidative type I1 a or I1 x fibers. Both muscles were identified as mainly slow and fatigue resistant.
Cytogenetic Results In 20.5 % of the examined metaphases (41 of 200 cells), cytogenetic alterations were observed. In about a half of these altered cells (18 cells) a small marker chromosome was observed (Fig. 4 a, b, c, d and Fig. 5 ) in addition to the bovine diploid chromosome set (2 n = 60). Thus, 61 chromosomes were present in these cells. Moreover, chromosome breakages and fragments predominated among the cytogenetic alterations (Fig. 4 e and f). No such alterations were oberserved in simultaneously performed control cultures. From the 5 cows which had been inseminated, 3 became pregnant. Ectopia cordis was not observed in any of the newborns. Two calves had a normal clinical picture. One calf died 1 day post partum from unknown causes. Discussion Reports of ectopia cordis in man originate in 1671 (22), however, the first case in veterinary literature was published in 1818 (2). Since then, numerous descriptions have been made concerning morphologic abnormalities in calves with ectopia cordis. Whereas the sternal and abdominal types are found in man (24, dog ( 5 ) , pig (6), and pigeon (19), in cattle the lower cervical type seems to be most common (2, 8, 11, 12, 14, IS), although the sternal type has already been described (17). The percentage of total congenital defects which are cardiac in cattle, varies from 1 . 5 % to 9.6% and depends on breed and geographic region. For example, 1.5 '/o to 2.7 % has been reported in Germany (16, 23), whereas 1.9 YO in Kansas, USA (9), 8.3 Yo in Japan (lo), and 9.6 % in Switzerland (17) have been described. According to different authors, ectopia cordis accounts for 1 % to 45 YO of all cardiac defects in cattle: 1 % (17), 13 % (S), 15 O/O (16) and 45 % (9). Surgical correction has been tried in man (22) and cattle (I), but has been entirely unsuccessful. We describe a calf with ectopia cordis cervicalis inferior with certain anatomic abnormalities of the great vessels which result in several hemodynamic alterations. To our knowledge this is the first description of functional properties of a congenital extrathoracic heart. The position of the great vessels is comparable to that of a bovine fetus 35 days after conception (7). At that time, the embryonic heart is near the 5.-6. cervical segment and the aortic arch near the 6. segment. Due to the lack of fusion of both cranial caval veins, the V. cava man. sin. remained. The V.jugularis externa, subclavia, and azygos drain into the Vv. cavae cran. of the corresponding side of the body. The persistent aortic arch at the 6. cervical segment prevents the junction of the Tr. brachiocephalicus with the A . subclavid sin. Similar morphologic observations such as persistence of the left cranial caval vein, emptying of one common pulmonary vein into an elongated left atrium, changes in the course of the arteries, and the sharp ventral bend of the sternum have been reported previously (2, 11, 12, 18). In our study we observed massive concentric myocardial hypertrophy. The cause of the left ventricular hypertrophy is unclear and may be multifactorial. It seems possible that the abnormal course of three thin arteries, both carotid arteries and one subclavian artery, arising from an enlarged Tr. brachiocephalicus, and the rightangled aortic arch result in an elevated afterload. Furthermore, the arterial hypertension increases afterload. Both the left ventricular pressure and the arterial pressure were elevated, although information on the exact value can not be provided due to the exaggerated movement of the heart within loose connective tissue. The dp/dt of the LVP was estimated in some circumstances as being up to 8000 torr/ sec whereas most of the time normal values were measured. The irregular cardiac rhythm together with the elevated heart rate and the increased LVP would contribute to this increased dp/dt during several cardiac cycles. Following the retraction of the catheter from
Cardiac Function, Morphology and Chromosomal Aberrations in a Calf
767
the left ventricle into the aorta a gradient of about 40 mm Hg was observed. Based on the observation that both the length of the aorta and its diameter, at least in the Sinus valsaluae and after the right angled aortic arch, was increased in this animal (see Fig. 2), an increase of the aortic compliance may be presumed. This increased compliance may also explain the low pulsatility of the aortic pressure. Furthermore, functional aortic stenosis following luxation of the heart, swinging during the cardiac cycles, might also contribute to the observed effect. Nevertheless, the LVP as well as the peripheral arterial pressure were elevated. The etiology of the hypertension is unclear. It may be an essential hypertension or may be due to altered baroreceptor sensitivity secondary to the abnormal great vessel anatomy, such as it can be seen in aortic coarctation. The hypertrophy of the right ventricle may indicate increased pulmonary vasculary resistance. Unfortunately, direct hemodynamic measurements in the pulmonary circulation could not be performed due to the kinking of the left cranial caval vein. However, since both ventricles showed myocarditis simplex, a primary muscle disease cannot be excluded. We observed that the resting calf appeared clinically normal, with normal pulse and color of the mucous membranes, whereas in a state of increased metabolic demand, even trotting, the animal showed signs of cardiac decompensation such as increase of the heart rate, cyanosis and dyspnoe. However, the function of liver and kidney was maintained. Cytogenetically, a major fraction of the investigated cells (mosaicism) of the calf revealed the presence of an additional small marker chromosome, giving rise to a chromosome number of 61. This marker possesses a clearly recognizable centromer. Moreover, the presence of two G-bands was typical for this small marker. However, a more definite cytogenetic characterisation, including an unequivocal identification, was not possible. Of further interest were the breakages observed in many metaphases. Chromosome breakage syndromes with background values of 1.8 % and 2 %, found in cattle (20, own unpublished data), play some role in several hereditary diseases at least in man. In our extopia calf, we observed a value of 20.5% of alterations which is clearly increased. Different chromosomes, in most cases hardly to be identified, were affected. High percentages of alterations (12 %, 15 Yo) have also been observed in two cases of bovine congenital harilessness (21). Similar to our present case, the increased alteration frequency could not be traced back to a certain chromosome segment. The 3 clinically normal offspring of our bullock showed a normal thoracic radiogram, in agreement with a previous report on normal offspring produced from a cow with ectopia cordis cervicalis (2). We conclude that in this case of ectopia cordis cervicalis inferior, the changed anatomic situation in the great vessels possibly led to increased afterload resulting in ventricular hypertrophy and compensated cardiac failure. Peripheral hypertension, either essential or as a result of altered baroreceptor activity, was also observed. Except for the markedly altered anatomic situation, the cardiac output was maintained at rest. Acknowledgement This study was supported by the Ludwig-Boltzmann Foundation. References 1. ASLANBEY, D., M.K. GAVUZOGLU, T.T~ssas,S.BERKIN, and B.YILMAZ, 1980: Bir danada ectopia cordis cervicalis olgusu. Veteriner Facultesi Dergisi Ankara Universitesi 27, 613 -632. 2. BOWEN, J.M., and R. W. ADRIAN, 1962: Ectopia cordis in cattle. J. Am. Vet. Med. Assoc. 141, 1162- 1167. 3. BROOKE, M. H., and K. K. KAISER,1970: Muscle fibre types: how many and what kind? Arch. Neurol. 23, 369-379. 4. BRUMBACK, R. A., and R. W. LEECH,1984: Color atlas of muscle histochemistry. Littleton, Massachusetts: PSG Publishing Company Inc., pg. 16.
768
URSULAWINDBERGER et al.
5. FREEMAN, L. E., and P. T. MCGOVERN, 1984: Thoracoabdominal ectopia cordis in two littermate pups. Canine Practice 11, 39-43. L. E., and P.T. MCGOVERN, 1984: Ectopia cordis thoracoabdominalis in a piglet. Vet. 6. FREEMAN, Rec. 115, 431-433. 7. GOLUB,D. M., 1931: Die Entwicklung des Aortenbogens und der von ihm ausgehenden Aste bei Saugetieren. I. Entwicklung der Kiemenbogen bei Kalb und Menschen. Zschr. Anat. Entw. Gesch. 95, 447-472. 8. GOPAL,T., H. W. LEIPOLD,and S. M. DENNIS, 1986: Congenital cardiac defects in calves. Am. J. Vet. Res. 47, 1120-1121. J. L. NOORDSY, and S. M. DENNIS,1973: Congenital 9. GREENE,H. J., H . W. LEIPOLD,K. HUSTON, defects in cattle. Irish Vet. J. 27, 37-45. 10. HAGIO,M., 1985: Congenital heart disease in cattle. Bull. Fac. Agric. Miyazaki Univ. 32, 233-250. 11. HIRAGA, T., and M. ABE, 1986: Eight calves of cervical ectopia cordis and their sternums. Jpn. J. Vet. Sci. 48, 1199- 1206. 12. HUGHES,H.V., 1934: Ectopia cordis in the calf. J. Comp. Path. Ther. 47, 141-151. 13. JAKSCH,W., und E. GLAWISCHNIG, 1981: Klinische Propadeutik der Inneren Krankheiten und Hautkrankheiten der Haustiere. Paul Parey Scientific Publishers, Berlin and Hamburg, 2nd edition. 14. JANIAK, T., 1954: Wrodzone przemieszczenie serca na szyje u rocznego buhaja. Med. Wtryn. 10, 83 - 86. 15. KANEKO, J. J. (ed.), 1980: Clinical biochemistry of domestic animals. Academic Press, New York - London - Toronto - Sydney - SanFrancisco, 3rd edition. H. H., 1985: Statistische Analyse des Auftretens von Kalbern mit kongenitalen 16. KOMOROWSKY, Anomalien in der Nachkommenschaft von Besamungsbullen in Hessen, eine explorative Datenbeschreibung. Dissertation Justus-Liebig-Univ. GieBen. und R. FATZER,1980: Angeborene morphologische Anomalien bei Kalbern 17. KONIG,B., A. TONTIS aus dem Raum Bern. Schweiz. Arch. Tierheilk. 122, 435-458. 18. KULCZYCKI, W., 1902: Ein Fall von Ectopia cordis beim Kalbe. Polnisches Archiv fur biologische und medizinische Wissenschaften 1, 1 - 15. and K. MEYER,1983: Ectopia cordis bei einer Taube. Dtsch. 19. KUMMERFELD, N., U. NEUMANN, tierarztl. Wschr. 90, 179- 180. 20. LOJDA, L., J. RUBES,and J. MRAZ, 1984: Levels of spontaneous chromosome aberrations in populations of cattle and pigs in Czechoslovakia as indicators of environmental mutagenic effects. Proceedings of the 6th European Colloquium o n Cytogenetics of Domestic Animals, Zurich, 104-106. 21. MAYR, B., 1979: Untersuchungen zytogenetischer Marker und ihrer Anwendbarkeit in der Zuchtpraxis bei Rind und Schwein. Habilitationsschrift, Veterinarmed. Universitat Wien. 22. MILLHOUSE, R. F., and H.A. JOOS, 1959: Extrathoracic ectopia cordis: report of cases and review of literature. Am. heart J. 57, 470-476. 1972: Untersuchungen zur teratologischen 23. RIECK, G. W., K. H. FINGER und A. HERZOG, Populationsstatistik und 7ur Atiologie der embryonakn Entwicklungsstorungen beim Rind. 1. Teratologische Populationsstatistik. GieDener Beitr. Erbpath. Zuchthyg. 4, 39-69. 24. WANG,H.C., and S.FEDOROFF,1972: Banding in human chromosomes treated with trypsin. Nature New Biol. 235, 52-54.
