REVIEW URRENT C OPINION
What does morphokinetics add to embryo selection and in-vitro fertilization outcomes? Natalia Basile a, Mauro Caiazzo a, and Marcos Meseguer b
Purpose of review Time lapse technology represents a new tool in the in-vitro fertilization (IVF) laboratory. It can aid the embryologist in the detection of objective and quantifiable markers associated with embryo viability and implantation. The purpose of this review is to explain how embryo morphokinetics can be used as an adjunct to standard morphological assessment and to evaluate its potential value to improve IVF outcomes. Recent findings Several algorithms have been developed. Some utilize early kinetic markers, whereas others rely more on later stages of embryo development. Even though over a handful of randomized control trials are in progress, at this point, only one has been published demonstrating a significant increase in implantation rates and ongoing pregnancy rates when selecting embryos on the basis of a combination of morphological assessment and morphokinetics. Summary We believe that standard morphological assessment should remain the gold standard to initiate embryo evaluation; however, if possible, it can be complemented with the use of morphokinetics. This new approach will allow the embryologist to perform a more accurate and objective embryo selection and therefore reduce the number of embryos transferred while maintaining or even improving clinical results. Keywords implantation, morphokinetics, randomized controlled trial
INTRODUCTION The study of embryo morphokinetics has resulted in the identification of different kinetic markers associated with embryo viability, implantation and live birth rates [1–5]. Some of these markers are being used to ‘select’ embryos, although others have turned useful to ‘de-select’ embryos. This new tool is of particular interest especially in Europe, where legislation regarding the number of embryos transferred is stricter, obliging in-vitro fertilization (IVF) practitioners to reduce the number of embryos transferred while maintaining good clinical results. As a consequence, the availability of time-lapse systems (TMS) is increasing allowing the users to choose according to their own needs and demands. There are many advantages that we could mention regarding TMS: maintenance of optimal culture conditions, continuous monitoring, detection of abnormal events that would normally occur between observations, minimization of culture media, gas and oil, reduced number of hours needed by the embryologist within the laboratory and so on. However, the question still remains: has the introduction of TMS added any value to the process
of embryo selection and IVF outcomes? We believe the answer is yes and the intention of this review is to justify such answer. To this aim, we will briefly review the standard methods of embryo evaluation to further continue with the description of novel morphokinetic markers. Finally, we will summarize the most relevant studies that justify the use of this new strategy for embryo selection.
VINTAGE EMBRYO ASSESSMENT: STATIC MORPHOLOGY At the moment, standard morphological assessment remains the most common method to evaluate embryo quality. Even though other noninvasive evaluation methods are being developed and constantly improved, morphology continues to be the a
IVI Madrid, Madrid and bIVI Valencia, Valencia, Spain
Correspondence to Natalia Basile, PhD, IVI Madrid, Av. Del Talgo 68, 28023 Madrid, Spain. Tel: +34 91 1802900; fax: +34 91 1802910; e-mail: [email protected] Curr Opin Obstet Gynecol 2015, 27:193–200 DOI:10.1097/GCO.0000000000000166
1040-872X Copyright ß 2015 Wolters Kluwer Health, Inc. All rights reserved.
www.co-obgyn.com
Copyright © 2015 Wolters Kluwer Health, Inc. All rights reserved.
Fertility, IVF and reproductive genetics
KEY POINTS The use of kinetic markers as a complement to standard morphological assessment improves embryo selection by the embryologist. Validation of this new strategy has been performed through a RCT demonstrating significant increase in implantation and ongoing pregnancy rates when it was utilized. Automatization has been crucial for the implementation of this technology on a clinical setting.
most widespread and effective system in use. However, concerns about the effects of handling the embryos outside stable culture conditions have limited the frequency of microscopic observations. One of the key disadvantages of evaluating embryos according only to morphological parameters is that the assessment is not always performed at the same time points, and these variations can be relevant. For instance, the same day-2 embryo may be at the two-cell stage early on day 2 or already at the fourcell stage if assessed later. Furthermore, the morphological appearance of an embryo can change rapidly, which may be relevant for day-2 and day-3 evaluations and can result in a classification that is not objectively correct [6]. The evaluation of different morphological criteria at different embryo stages has been studied in order to correlate them with the probability of successful embryo implantation [7–13]. Evaluation at these stages has traditionally formed the basis for determining embryo quality. However, although there is a certain amount of agreement on what constitutes a good or poor quality embryo, it is very difficult to find a consensus between laboratories when attempting to evaluate intermediate quality embryos. Moreover, the subjectivity in embryo quality can result in highly variable embryo scores depending on the embryologist [14]. To overcome this issue, embryologists from ALPHA and the European Society of Human Reproduction and Embryology (ESHRE) have elaborated a consensus describing the minimum data set required for the accurate description of embryo development (Istanbul Consensus, 2011). The purpose of this consensus was not only to define common terminology but also to standardize laboratory practice in order to allow for more effective comparisons of treatment outcomes between different groups [15]. It is not our intention to describe this consensus in detail; however, a brief summary is presented in Tables 1 and 2. According 194
www.co-obgyn.com
to the consensus, the timing of observations is critical and should be standard between different laboratories. In addition, it was noted that for embryos, each observation should have two parts: cell number/stage and grading. Interestingly, this consensus comments that more frequent or prolonged observations of embryos (in addition to the minimum set established) carry the risk (albeit small) of an impact in their development. The introduction of TMS overcomes this issue and it is now possible to perform continuous observations of embryo development with no risk at all [16,17]. This constitutes one of the first advantages with respect to standard incubation.
MODERN EMBRYO ASSESSMENT: DYNAMIC MORPHOKINETICS The modern era for embryo quality assessment takes a different approach: dynamic morphokinetics. This new concept is the result of the recent implementation of TMS in the IVF laboratories and it combines the concept of embryo morphology with the kinetics of embryo development.
KINETIC PARAMETERS According to the proposed guidelines on the nomenclature and annotation of dynamic human embryo monitoring [18 ], we can define the following morphokinetic ‘individual’ parameters (supplemental video 1, http://links.lww.com/COOG/A19). Figure 1 describes some of them. &&
(1) t0
Time of IVF or mid-time of micro/injection (ICSI/IMSI)
(2) tPB2
The second polar body completely detached from the oolemma
(3) tPN
Fertilization is confirmed
(4) tPNa
Appearance of individual pronuclei; tPN1a, tPN2a; tPN3a and so on
(5) tPNf
Time of pronuclei disappearance; tPN1f; tPN2f; and so on
(6) tZ
Time of pronuclei scoring
(7) t2–t9
Time to two to nine discrete cells
(8) tSC
First evidence of compaction
(9) tMf/p
End of compaction ‘f’, fully compacted ‘p’, partial compaction
(10) tSB
Initiation of blastulation
(11) tByz
Full blastocyst ‘y’, morphology of inner cell mass ‘z’, morphology of trophectoderm cells
(12) tEyz
Initiation of expansion; first frame of zona thinning
(13) tHNyz
Herniation; end of expansion phase and initiation of hatching
(14) tHDyz
Fully hatched blastocyst
Volume 27 Number 3 June 2015
Copyright © 2015 Wolters Kluwer Health, Inc. All rights reserved.
Rating
Good
Fair
Poor
Grade
1
2
3
Cleavage stage
1040-872X Copyright ß 2015 Wolters Kluwer Health, Inc. All rights reserved.
Cell size not stage-specific Evidence of multinucleation
Severe fragmentation (>25%)
No evidence of multinucleation
Stage-specific cell size for majority of cells
10–25% fragmentation
No multinucleation
Stage-specific cell size
What does morphokinetics add to embryo selection and in-vitro fertilization outcomes? Natalia Basile a, Mauro Caiazzo a, and Marcos Meseguer b
Purpose of review Time lapse technology represents a new tool in the in-vitro fertilization (IVF) laboratory. It can aid the embryologist in the detection of objective and quantifiable markers associated with embryo viability and implantation. The purpose of this review is to explain how embryo morphokinetics can be used as an adjunct to standard morphological assessment and to evaluate its potential value to improve IVF outcomes. Recent findings Several algorithms have been developed. Some utilize early kinetic markers, whereas others rely more on later stages of embryo development. Even though over a handful of randomized control trials are in progress, at this point, only one has been published demonstrating a significant increase in implantation rates and ongoing pregnancy rates when selecting embryos on the basis of a combination of morphological assessment and morphokinetics. Summary We believe that standard morphological assessment should remain the gold standard to initiate embryo evaluation; however, if possible, it can be complemented with the use of morphokinetics. This new approach will allow the embryologist to perform a more accurate and objective embryo selection and therefore reduce the number of embryos transferred while maintaining or even improving clinical results. Keywords implantation, morphokinetics, randomized controlled trial
INTRODUCTION The study of embryo morphokinetics has resulted in the identification of different kinetic markers associated with embryo viability, implantation and live birth rates [1–5]. Some of these markers are being used to ‘select’ embryos, although others have turned useful to ‘de-select’ embryos. This new tool is of particular interest especially in Europe, where legislation regarding the number of embryos transferred is stricter, obliging in-vitro fertilization (IVF) practitioners to reduce the number of embryos transferred while maintaining good clinical results. As a consequence, the availability of time-lapse systems (TMS) is increasing allowing the users to choose according to their own needs and demands. There are many advantages that we could mention regarding TMS: maintenance of optimal culture conditions, continuous monitoring, detection of abnormal events that would normally occur between observations, minimization of culture media, gas and oil, reduced number of hours needed by the embryologist within the laboratory and so on. However, the question still remains: has the introduction of TMS added any value to the process
of embryo selection and IVF outcomes? We believe the answer is yes and the intention of this review is to justify such answer. To this aim, we will briefly review the standard methods of embryo evaluation to further continue with the description of novel morphokinetic markers. Finally, we will summarize the most relevant studies that justify the use of this new strategy for embryo selection.
VINTAGE EMBRYO ASSESSMENT: STATIC MORPHOLOGY At the moment, standard morphological assessment remains the most common method to evaluate embryo quality. Even though other noninvasive evaluation methods are being developed and constantly improved, morphology continues to be the a
IVI Madrid, Madrid and bIVI Valencia, Valencia, Spain
Correspondence to Natalia Basile, PhD, IVI Madrid, Av. Del Talgo 68, 28023 Madrid, Spain. Tel: +34 91 1802900; fax: +34 91 1802910; e-mail: [email protected] Curr Opin Obstet Gynecol 2015, 27:193–200 DOI:10.1097/GCO.0000000000000166
1040-872X Copyright ß 2015 Wolters Kluwer Health, Inc. All rights reserved.
www.co-obgyn.com
Copyright © 2015 Wolters Kluwer Health, Inc. All rights reserved.
Fertility, IVF and reproductive genetics
KEY POINTS The use of kinetic markers as a complement to standard morphological assessment improves embryo selection by the embryologist. Validation of this new strategy has been performed through a RCT demonstrating significant increase in implantation and ongoing pregnancy rates when it was utilized. Automatization has been crucial for the implementation of this technology on a clinical setting.
most widespread and effective system in use. However, concerns about the effects of handling the embryos outside stable culture conditions have limited the frequency of microscopic observations. One of the key disadvantages of evaluating embryos according only to morphological parameters is that the assessment is not always performed at the same time points, and these variations can be relevant. For instance, the same day-2 embryo may be at the two-cell stage early on day 2 or already at the fourcell stage if assessed later. Furthermore, the morphological appearance of an embryo can change rapidly, which may be relevant for day-2 and day-3 evaluations and can result in a classification that is not objectively correct [6]. The evaluation of different morphological criteria at different embryo stages has been studied in order to correlate them with the probability of successful embryo implantation [7–13]. Evaluation at these stages has traditionally formed the basis for determining embryo quality. However, although there is a certain amount of agreement on what constitutes a good or poor quality embryo, it is very difficult to find a consensus between laboratories when attempting to evaluate intermediate quality embryos. Moreover, the subjectivity in embryo quality can result in highly variable embryo scores depending on the embryologist [14]. To overcome this issue, embryologists from ALPHA and the European Society of Human Reproduction and Embryology (ESHRE) have elaborated a consensus describing the minimum data set required for the accurate description of embryo development (Istanbul Consensus, 2011). The purpose of this consensus was not only to define common terminology but also to standardize laboratory practice in order to allow for more effective comparisons of treatment outcomes between different groups [15]. It is not our intention to describe this consensus in detail; however, a brief summary is presented in Tables 1 and 2. According 194
www.co-obgyn.com
to the consensus, the timing of observations is critical and should be standard between different laboratories. In addition, it was noted that for embryos, each observation should have two parts: cell number/stage and grading. Interestingly, this consensus comments that more frequent or prolonged observations of embryos (in addition to the minimum set established) carry the risk (albeit small) of an impact in their development. The introduction of TMS overcomes this issue and it is now possible to perform continuous observations of embryo development with no risk at all [16,17]. This constitutes one of the first advantages with respect to standard incubation.
MODERN EMBRYO ASSESSMENT: DYNAMIC MORPHOKINETICS The modern era for embryo quality assessment takes a different approach: dynamic morphokinetics. This new concept is the result of the recent implementation of TMS in the IVF laboratories and it combines the concept of embryo morphology with the kinetics of embryo development.
KINETIC PARAMETERS According to the proposed guidelines on the nomenclature and annotation of dynamic human embryo monitoring [18 ], we can define the following morphokinetic ‘individual’ parameters (supplemental video 1, http://links.lww.com/COOG/A19). Figure 1 describes some of them. &&
(1) t0
Time of IVF or mid-time of micro/injection (ICSI/IMSI)
(2) tPB2
The second polar body completely detached from the oolemma
(3) tPN
Fertilization is confirmed
(4) tPNa
Appearance of individual pronuclei; tPN1a, tPN2a; tPN3a and so on
(5) tPNf
Time of pronuclei disappearance; tPN1f; tPN2f; and so on
(6) tZ
Time of pronuclei scoring
(7) t2–t9
Time to two to nine discrete cells
(8) tSC
First evidence of compaction
(9) tMf/p
End of compaction ‘f’, fully compacted ‘p’, partial compaction
(10) tSB
Initiation of blastulation
(11) tByz
Full blastocyst ‘y’, morphology of inner cell mass ‘z’, morphology of trophectoderm cells
(12) tEyz
Initiation of expansion; first frame of zona thinning
(13) tHNyz
Herniation; end of expansion phase and initiation of hatching
(14) tHDyz
Fully hatched blastocyst
Volume 27 Number 3 June 2015
Copyright © 2015 Wolters Kluwer Health, Inc. All rights reserved.
Rating
Good
Fair
Poor
Grade
1
2
3
Cleavage stage
1040-872X Copyright ß 2015 Wolters Kluwer Health, Inc. All rights reserved.
Cell size not stage-specific Evidence of multinucleation
Severe fragmentation (>25%)
No evidence of multinucleation
Stage-specific cell size for majority of cells
10–25% fragmentation
No multinucleation
Stage-specific cell size
![[Does embryo morphology constitute a reliable criterion for embryo selection?].](/assets/img/hold.png)
