Human embryonic cardiovascular function

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1 AOGS REVIEW ARTICLE Human embryonic cardiovascular function GANESH ACHARYA 1,2,3, YONGHAO GUI 4, WOJCIECH CNOTA 5, JAMES HUHTA 6 & AGATA WLOCH 5 1 Women s Health and Perinatology Research Group, Department of Clinical Medicine, Faculty of Health Sciences, UiT The Arctic University of Norway, Tromsø, 2 Department of Obstetrics and Gynecology, University Hospital of Northern Norway, Tromsø, Norway, 3 Department of Clinical Sciences, Intervention and Technology, Karolinska Institute, Stockholm, Sweden, 4 Cardiovascular Center, Fudan University Children s Hospital, Shanghai, China, 5 Clinical Department of Obstetrics and Gynecology, Chair of Women s Health, School of Health Sciences, Medical University of Silesia, Katowice, Poland, and 6 Perinatal Cardiology, All Children s Hospital, Pediatrix Medical Group, St Petersburg, Florida, USA Key words Human embryo, Doppler, cardiac function, embryonic heart, embryonic circulation Correspondence Ganesh Acharya, Department of Obstetrics and Gynecology,University Hospital of Northern Norway, Sykehusveien 38, 9038 Tromsø, Norway. ganesh.acharya@uit.no Conflicts of interest The authors have stated explicitly that there are no conflicts of interest in connection with this article. Please cite this article as: Acharya G, Gui Y, Cnota W, Huhta J, Wloch A. Human embryonic cardiovascular function. Acta Obstet Gynecol Scand 2016; 95: Received: 10 September 2015 Accepted: 18 January 2016 DOI: /aogs Abstract Introduction. This review presents an overview of descriptive knowledge on human embryonic cardiovascular physiology mostly based on noninvasive assessment by Doppler ultrasonography. Our objective was to identify and analyze published studies on embryonic cardiovascular function, and summarize available knowledge in this field. Material and methods. Citations related to human embryonic cardiovascular function were searched in PubMed, EMBASE, CINAHL and Web of Science using keywords and MeSH terms without any time limitation. The search was restricted to English language articles. Abstracts were screened and full texts of relevant articles were obtained. All articles that reported on physiological aspects of human embryonic cardiovascular function were included. Studies reporting on cardiovascular function after 10 weeks of gestation were excluded. Data were synthesized and presented narratively. Results. We identified 10 studies that had evaluated cardiovascular function and/or hemodynamics in human embryos at 10 weeks of gestation. All of these reported only certain aspects of embryonic cardiovascular function. Embryonic heart rate is associated significantly with gestational age and increases from 6 to 10 weeks of gestation. Cardiac inflow is monophasic during the embryonic period and atria appear to generate higher force during contraction compared with ventricles. Both ventricular inflow and outflow velocities increase with advancing gestation, whereas the Tei index decreases significantly. During the embryonic period, placental blood flow increases with gestation, but absent umbilical artery diastolic flow and umbilical venous pulsations are normal phenomena. Conclusion. There are important differences in normal cardiovascular function between the embryonic and fetal stages of human in utero development. Abbreviations: ICT, isovolumic contraction time; IRT, isovolumic relaxation time. Introduction The cardiovascular system is one of the first organ systems to develop in humans in utero (1). When describing the time sequence of embryonic development, it is important to remember that the postconceptional gestational age usually used by embryologists differs from the postmenstrual gestational age generally used in clinical practice. In women with regular menstrual cycles of 28 days, ovulation (and therefore presumed conception) occurs approximately 2 weeks before the next expected menstruation. Therefore, the embryo is about 2 weeks younger than the clinically determined gestational age. ª 2016 Nordic Federation of Societies of Obstetrics and Gynecology, Acta Obstetricia et Gynecologica Scandinavica 95 (2016)

2 Embryonic cardiac function G. Acharya et al. Postmenstrual age is less accurate because of the varying length of menstrual cycles, especially in women with irregular cycles. Accurate determination of the date of fertilization (conception) is difficult unless the pregnancy was conceived by in vitro fertilization or using other assisted reproductive techniques. Therefore, ultrasoundmeasured crown rump length is generally used to date the pregnancy in the first trimester, as this method has been found to be reasonably accurate (2). For the purpose of this review a crown rump length 30 mm, which corresponds to 10 weeks of gestation (3), is considered as the embryonic period, and the gestational age refers to postmenstrual gestational age unless specified otherwise. The human embryonic heart develops during the 3rd and 8th weeks after conception (1), and its structures are well defined by the end of the embryonic period. Abnormal development during this period of organogenesis is mostly responsible for congenital heart defects, which are the most common congenital malformations and the leading cause of infant mortality in developed countries. They may also lead to embryonic/fetal demise. If abnormalities of the developing heart could be detected during the embryonic period, it could help to predict pregnancy outcome. Interventions applied early during development might even help to ameliorate or prevent functional damage. Echocardiography is a well-established method of evaluating fetal heart structure and function. As organogenesis is complete by 10 weeks of gestation and the developing heart has reached its definitive anatomic form for intrauterine life, echocardiography with high-frequency high-resolution probes, can be used to visualize structures of the fetal heart in the late first trimester. This helps to confirm normality, detect certain congenital defects and evaluate cardiac function. The four-chamber view is possible to obtain in more than 90% of examinations from 12 weeks onward, and the ability to perform a full cardiac examination increases from about 20% in week 11 to 92% in week 13 (4). However, it is almost impossible to diagnose a structural heart defect in the embryonic period because of the small cardiac size and rapidly changing morphology. Furthermore, during the period of organogenesis, the embryonic heart undergoes complex morphological and physiological changes, which are reflected in the variability of measured parameters used to assess its function. The anatomical growth and functional maturity of the cardiovascular system in utero continue beyond the embryonic period. Although it is challenging to perform echocardiography during the embryonic period (<10 weeks), some aspects of cardiovascular function of the developing embryonic heart can be studied using Doppler as early as 6 weeks of gestation (5 10). As cardiac structure is closely associated with function, abnormal cardiac function could indicate abnormal heart development (11). However, the lack of sufficient data defining normality is a major impediment in diagnosing functional abnormalities during the embryonic period. The aim of this review was to identify and analyze published studies on embryonic cardiovascular function, and summarize available knowledge in this field. Material and methods Citations on human embryonic cardiovascular function were searched in PubMed, EMBASE, CINAHL and Web of Science using MeSH terms and keywords (embryo, physiology, Blood supply, circulation, heart, cardiac function, Doppler, echocardiography). The search was restricted to English language articles, but without any restriction for publication date. Reference lists of eligible studies were manually searched. Abstracts were screened and full texts of all relevant articles were obtained. All articles that reported on physiological aspects of human embryonic cardiovascular function were included. Studies reporting on cardiovascular function after 10 weeks of gestation were excluded. Two investigators (GA and AW) independently evaluated the articles for methodological quality using the National Institutes of Health Quality Assessment Tool for Observational Cohort and Cross-sectional Studies, extracted data, and created a summary table with key characteristics of all included studies (Table 1). Any disagreement was resolved by mutual discussion leading to consensus. Data were synthesized and presented narratively. Results We identified 10 studies that had evaluated cardiovascular function and/or hemodynamics in human embryos <10 weeks of gestation. All of them were of fair quality. Five studies (5 9) were longitudinal, and five (10,12 15) had a cross-sectional design. All articles reported only certain aspects of embryonic cardiovascular function, and Key Message Significant changes in cardiovascular function are observed from the start of cardiac activity at 5 weeks of gestation to the end of the embryonic period (10 weeks) in the developing human embryo. There are important differences in normal cardiovascular function between the embryonic and fetal stages of in utero development. 622 ª 2016 Nordic Federation of Societies of Obstetrics and Gynecology, Acta Obstetricia et Gynecologica Scandinavica 95 (2016)

3 G. Acharya et al. Embryonic cardiac function Table 1. Characteristics of the studies included in the review. Studies included Type of study Risk group Number of embryos GA, weeks Embryonic cardiac function parameters Embryonic peripheral flow parameters Arduini et al., 1991 (13) Rizzo et al., 1992 (12) Van Splunder et al., 1996 (14) Merce et al., 1996 (15) Leiva et al., 1999 (5) M akikallio et al., 1999 (7) M akikallio et al., 2005 (6) Wloch et al., 2007 (8) Wloch et al., 2008 (9) Wloch et al., 2015 (10) C/S prospective LR UA PI C/S prospective LR 99 (257) 7 10 (7 16) HR UA PI, UV waveform, TVI of IVC C/S prospective LR Blood flow velocity waveform of UA, UV, DV, IVC, and DAo C/S prospective LR UA PI, UV waveform, Uteroplacental, retrochorionic and intervillous circulation L/S prospective HR (IVF HR, HS, ET, FT, group) ICT, IRT, valve clicks L/S prospective LR HR Blood flow velocity waveforms of yolk sac, UA and chorionic arteries L/S prospective LR HR, ICT, IRT, outflow V max, and V mean, Inflow waveform L/S prospective LR HR, HS, ICT, IRT, ET, MPI, outflow V max and V mean, Inflow waveform UA, UV and DV blood flow velocity waveform analysis UA, UV and DV blood flow velocity waveform analysis L/S prospective LR HR UA, UV and DV blood flow velocity waveform analysis C/S prospective LR V mean inflow, V mean outflow EF inflow /EF outflow ratio C/S, cross sectional; LR, low risk; HR, high risk; GA, gestational age; HR, heart rate; HS, heart size; ICT, isovolumic contraction time; IRT, isovolumic relaxation time; ET, ejection time; FT, filling time; MPI, myocardial performance index; DV, ductus venosus; UA, umbilical artery; PI, pulsatility index; V mean, mean velocity; V max, maximum velocity; IVC, inferior vena cava; DAo descending aorta. a few attempted to correlate abnormal cardiovascular function with pregnancy outcome. Cardiac function The human embryonic heart is a spontaneously contracting cylindrical tube at approximately 5 weeks of gestation (21 days after conception) (16). It subsequently undergoes a series of changes, such as looping, septation, valve formation and compaction. The tubular heart appears to obliterate its lumen completely during systole (i.e. has a 100% ejection fraction) as demonstrated by high-resolution M-mode echocardiography (17) and optical coherence tomography in chick embryos (18). The cardiac motion can be visualized and recorded using B-mode and M-mode echocardiography after 5 weeks of gestation. The heart almost triples its size (biventricular diameter) from mm to mm during 6 10 weeks of gestation (8). Cardiomegaly is a cardinal sign of heart failure, and cardiac size (biventricular diameter, cardiac circumference or area) in relation to thorax size is often used in the assessment of fetal cardiac function (19). However, whether embryonic heart failure is associated with cardiomegaly is not known. Although primitive blood starts to circulate through the embryo quite early in gestation, the spatio-temporal resolution of echocardiography is not sufficient to visualize and record the human embryonic circulation before 6 weeks of gestation. The Doppler cardiac flow velocity waveform patterns obtained during the earliest stages of human embryonic development are similar to those observed in the embryonic mouse (20). Altered hemodynamics can affect early as well as later stages of cardiovascular development. Parameters describing the embryonic heart function differ from those used to describe fetal heart function later in pregnancy (19,21). It is not possible to calculate volumetric flows, such as stroke volume and cardiac output, using the measurements of flow velocities and valve diameters, as the cardiac valves are not yet completely developed and their cross-sectional areas are too small to measure accurately. Calculation of the ventricular shortening fraction using ª 2016 Nordic Federation of Societies of Obstetrics and Gynecology, Acta Obstetricia et Gynecologica Scandinavica 95 (2016)

4 Embryonic cardiac function G. Acharya et al. M-mode echocardiography is also not possible because of difficulties associated with identifying the cardiac chambers before septation is complete. Doppler technique is useful in the assessment of embryonic cardiovascular function as flow velocity waveforms with distinctly recognizable patterns can be recorded from the heart and peripheral circulation. The usually measured parameters include cardiac cycle time intervals, ventricular inflow and outflow velocities and Doppler blood flow velocity waveforms of the ductus venosus, umbilical artery and umbilical vein. Heart rhythm and rate After 5 weeks of gestation the heart rhythm is generally regular and the ventricular contraction is normally preceded by an atrial contraction. Gestational-age-associated changes in heart rate during the embryonic period have been described by several investigators (8,22 25) and a mean heart rate of about bpm at 6 weeks of gestation increasing to bpm at 10 weeks of gestation has been reported in most studies. The association between embryonic heart rate and the gestational age is quite strong and it has even been suggested as an alternative method for dating pregnancy (26). The M-mode technique is preferable for the evaluation of embryonic heart rate in clinical settings because of its high temporal resolution and better safety compared with pulsed-wave Doppler. However, both techniques have been used. Abnormal heart rate during 6 8 weeks of gestation is associated with poor pregnancy outcome (27). An embryonic heart rate of <80 90 bpm carries a dismal prognosis with a very high likelihood of miscarriage before the end of the first trimester (28,29). Furthermore, embryos with slow heart rates seem to have a greater risk of having anomalies than embryos with normal heart rates (30). Atrial function In the embryonic human heart, the ventricular inflow is monophasic, and ventricular filling seems to be dependent on the atrial contraction (5,8). Atrial dominance in the embryonic period of heart development has been recently described. The force generated by the atria during contraction appears to be higher than that generated by the ventricles (10). Valvular function Movements of the valves during their opening and closure can be recorded as strong echogenic signals just before the start and after the cessation of blood flow, while recording the pulsed-wave Doppler velocity waveforms of the atrioventricular and semilunar valves. They are referred to as valve clicks. Leiva et al. (5) first reported the timing of the onset of atrioventricular and semilunar valve clicks in normal human embryos. In a longitudinal study, Wloch et al. (8) were not able to demonstrate valve clicks before 8 weeks of gestation and the semilunar valve clicks appeared for the first time at 9 weeks of gestation in 30% of embryos. At 10 weeks, 80% of embryos examined had semilunar valve clicks (8). M akikallio et al. observed valve clicks at 7 weeks of gestation in 10% of embryos examined (6). Semilunar valves are generally competent and leakage does not normally occur in developing human embryos. However, there appears to be some disagreement regarding the competency of atrioventricular valves during the embryonic period. M akikallio et al. have reported nonholosystolic atrioventricular valve regurgitation in one out of 16 embryos examined before 9 weeks of gestation and in four out of 16 (25%) during 9 10 weeks of gestation (6). However, atrioventricular valve regurgitation was not reported by Leiva et al. (5) in their study of embryos conceived after in vitro fertilization, and Wloch et al. (8) also could not find it in a much larger number of embryos examined before 10 weeks. Ventricular function The inflow outflow velocities and the time intervals of the different phases of cardiac cycle measured using pulsed-wave Doppler are used to describe embryonic ventricular function. During the embryonic period the cardiac ventricles seem less compliant as their filling is dependent on atrial contraction and the inflow velocity waveform is monophasic. Biphasic filling pattern appears first at 10 weeks of gestation. The isovolumic relaxation time (IRT) of the cardiac cycle has been used for the assessment of diastolic cardiac function, especially the early part of diastole. The earliest study that reported IRT of the embryonic heart found that it amounted to 16% of the cardiac cycle throughout the embryonic period (5). However, later studies have reported that the IRT % decreases significantly with advancing gestation (6,8), and it is not directly associated with ventricular filling pattern (6). The inflow is monophasic during the embryonic period and the maximum inflow velocities increase with gestation (from at 6 weeks to cm/s at 10 weeks). During the period between 7 and 10 weeks of gestation the maximum velocity of inflow is consistently higher than that of the outflow (8). Isovolumic contraction time (ICT) is a parameter used to assess systolic heart function. It is closely associated with gestational age. The ICT% decreases from 624 ª 2016 Nordic Federation of Societies of Obstetrics and Gynecology, Acta Obstetricia et Gynecologica Scandinavica 95 (2016)

5 G. Acharya et al. Embryonic cardiac function % of the cardiac cycle at 8 weeks to % at 9 weeks (p < ) (8). Similar observations were published in two other reports (5,6). There is a significant increase in maximum outflow velocities from to cm/s during 6 10 weeks of gestation (8). The Tei Index, i.e. (IRT + ICT)/Ejection time, is used to describe global (combined systolic and diastolic) ventricular function. In the late first trimester (11 14 weeks), the mean Tei index of the left ventricle is and that of the right ventricle is , and does not vary significantly with crown rump length (31). The Tei index values are much higher in the embryonic period. The Tei index decreases significantly from a mean of at 6 weeks to at 10 weeks (8). Arterial and venous blood flow In humans, the chorionic villi and their vasculature start developing after the 4th week of gestation. The first signs of vasculogenesis are observed at 5 weeks. At 6 weeks the vitelline (yolk sac) and umbilical (placental) circulations are connected to the primitive heart and fully functional. During 6 10 weeks, the blood flow velocities and the pulsatility index in the vitelline arteries do not change significantly (7). After the embryonic period, the yolk sac and vitelline blood flow regress. Absent diastolic blood flow in the umbilical artery (6,9,13 15) and descending aorta (14) is a typical finding during the embryonic period. However, the umbilical artery peak systolic velocities increase significantly during the embryonic period (15) and mean velocities increase more than threefold during 6 10 weeks of gestation without a significant change in the pulsatility index (a surrogate for placental impedance), suggesting that there is a gestational age associated increase in the placental volume blood flow (6). Ductus venosus blood velocity waveforms demonstrate an antegrade flow during the whole cardiac cycle (6,9) and the velocities do not change significantly during 6 10 weeks of gestation. Pulsations are typically present in the umbilical vein Doppler waveforms during the embryonic period (9,12,14,15). Typical findings observed on ultrasonographic examination of the cardiovascular system during the embryonic period (before 10 weeks of gestation) are demonstrated in Figure 1. Discussion A detailed noninvasive evaluation of structures and function of the developing human embryonic heart is difficult. There are limitations related to access and safety. Depth required to penetrate by the ultrasound beam for scanning the human embryo does not allow the use of Figure 1. Typical findings observed during ultrasonographic evaluation of embryonic cardiovascular system (<10 weeks of gestation). Upper left: M-mode echocardiography demonstrating heart motion. Upper right: Pulsed-wave Doppler recording of cardiac inflow and outflow demonstrating monophasic inflow (above the baseline) that has higher maximum velocity than the outflow (below the baseline). Lower left: Ductus venosus blood flow velocity waveforms demonstrating forward flow during the whole cardiac cycle. Lower right: Doppler recording of the umbilical cord demonstrating absent diastolic flow in the umbilical artery (above the baseline) and pulsations in the umbilical vein (below the baseline). ª 2016 Nordic Federation of Societies of Obstetrics and Gynecology, Acta Obstetricia et Gynecologica Scandinavica 95 (2016)

6 Embryonic cardiac function G. Acharya et al. very high frequency probes, and possible teratogenicity risk associated with exposure to high levels of mechanical and thermal energy cannot be ignored. Therefore, animal models remain useful for studying cardiac structures and function during normal and abnormal embryonic development. There are only a few studies that have investigated cardiac structural and/or hemodynamic changes that occur during the embryonic period of human development using B-mode and pulsed-wave Doppler ultrasonography and some normative data are available from 5 to 6 gestational weeks onwards. Bradycardia is an early prognostic marker of pregnancy loss during the embryonic period. An increased IRT% also appears to be associated with risk of fetal demise in the first trimester. Both of these perhaps occur because of myocardial dysfunction in this group of embryos (8). Another marker is the timing of occurrence of valvular clicks on inflow outflow Doppler recordings. Leiva et al. (5) observed that the valvular clicks were delayed in pregnancies that miscarried in a group of women who had undergone in vitro fertilization. However, due to the inherently high-risk nature of these pregnancies, it difficult to know whether hemodynamic data obtained from these embryos represent normal development (32). Further studies are needed to elucidate whether presence or absence of valve clicks at a particular gestation has any prognostic value. Discrepancy in reported findings regarding atrioventricular valve regurgitation among investigators is interesting. However, it is possible to confuse normal outflow tract blood velocity waveforms with atrioventricular valve regurgitation. Theoretically, one would expect the atrioventricular valve leakage to occur during the isovolumic contraction phase of the cardiac cycle immediately following the closure of the semilunar valves. However, between 7 and 9 weeks of gestation, M akikallio et al. (6) noted a delay of ms between the end of ventricular filling and the beginning of regurgitation, but not after 9 weeks. Furthermore, the interpretation of inflow and outflow velocity waveforms based on the direction of flow may not be valid during early developmental stages when the heart can be considered as a looping muscular tube. Therefore, the presence of non-holosystolic atrioventricular valve regurgitation in some embryos at 7 weeks of human pregnancy, as suggested by Makikallio et al. (6), remains doubtful because at that time cardiac cushions are not yet transformed into valves. Inflow maximum velocity from 9 weeks of gestation onwards exceeds the maximum velocity of outflow. This may be related to rise in the circulating blood volume in the embryo and increase in atrial contraction pressure. Developing heart requires specialized conduction tissue to allow atrioventricular synchrony at an early embryonic age along with contractile capability. With these two essential features in place, the embryonic heart appears to undergo dramatic changes in morphology including segmented tube formation, looping, septation, and valve development, while at the same time maintaining a progressive increase in forward flow without signs of valvular regurgitation. From correlative studies, we know that the specialization of tissue at the inflow and outflow of the developing ventricle allows perfect valvular function with no sign of either atrioventricular or semilunar valvular tissue. This pinching off tissue functioning as valves combined with the extraordinary atrial ejection force to assist ventricular filling succeeds in maintaining efficient ventricular ejection to the developing embryonic circulation during this crucial period. Ductus venosus pulsatility index is quite high during the embryonic period, ranging between 0.91 and 0.87 (9), but the flow is antegrade throughout the cardiac cycle (6,9). Normal antegrade blood flow in the ductus venosus but pulsations in the umbilical vein suggest that the pulsations are not due to the transmission of pressure waves during atrial contraction, but perhaps are related to other factors, such as transmission of arterial pulsations and low vascular compliance. Similarly, placental vascular impedance appears to be high during the whole embryonic period despite a significant increase in placental volume blood flow (6). Human embryonic cardiovascular function may be further elucidated in future as the imaging technology improves. However, safety remains a concern in embryonic examination using Doppler ultrasonography. A detailed description of issues related to ultrasound safety is beyond the scope of this review article. However, all the studies included in our review had ethical approval. Most recent studies have used the ALARA (As Low As Reasonably Achievable) principle and continuous on screen display of mechanical and thermal indexes keeping them below 0.7 and 0.5, respectively. Offline measurement of Doppler velocity waveforms and calculation of parameters describing cardiovascular function is recommended to reduce ultrasound exposure time. Conclusions Doppler ultrasonographic assessment of some aspects of human embryonic cardiovascular function is possible after 5 weeks of gestation. There are important differences in normal cardiovascular function between the embryonic and fetal stage of human in utero development. After 8 weeks of gestation, the heart is morphologically well formed, but has not yet achieved effective myocardial compliance. Throughout the embryonic period, the human 626 ª 2016 Nordic Federation of Societies of Obstetrics and Gynecology, Acta Obstetricia et Gynecologica Scandinavica 95 (2016)

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