scholarly journals Label-free assessment of pre-implantation embryo quality by the Fluorescence Lifetime Imaging Microscopy (FLIM)-phasor approach

2018 ◽  
Author(s):  
Ning Ma ◽  
Nabora Reyes de Mochel ◽  
Paula Duyen Anh Pham ◽  
Tae Yeon Yoo ◽  
Ken WY. Cho ◽  
...  
Keyword(s):  
Fluorescence Lifetime ◽  
Embryo Quality ◽  
Assisted Reproductive Technologies ◽  
Learning Algorithm ◽  
Reproductive Technologies ◽  
List Type ◽  
Label Free ◽  
Embryo Viability ◽  
Invasive Approach ◽  
Non Invasive

AbstractDevelopment of quantitative, safe and rapid techniques for assessing embryo quality provides significant advances in Assisted Reproductive Technologies (ART). We apply the phasor-FLIM method to capture endogenous fluorescent biomarkers of pre-implantation embryos as a non-morphological caliber for embryo quality. Here, we identify the developmental, or “D-trajectory”, that consists of fluorescence lifetime from different stages of mouse pre-implantation embryos. The D-trajectory correlates with intrinsic fluorescent species from a distinctive energy metabolism and oxidized lipids, as seen with Third Harmonic Generation (THG) that changes over time. In addition, we have defined an Embryo Viability Index (EVI) to distinguish pre-implantation embryo quality using the Distance Analysis, a machine learning algorithm to process the fluorescence lifetime distribution patterns. We show that the phasor-FLIM approach provides a much-needed non-invasive quantitative technology for identifying healthy embryos at the early compaction stage with 86% accuracy. This may increase embryo implantation success for in vitro fertilization clinics.HighlightsA label-free method of tracking metabolic trajectories during pre-implantation mouse embryo development.A non-invasive approach for assessing embryo quality and viability by a phasor-FLIM analysis.

scholarly journals Label-free assessment of pre-implantation embryo quality by the Fluorescence Lifetime Imaging Microscopy (FLIM)-phasor approach

2019 ◽  
Vol 9 (1) ◽  
Author(s):  
Ning Ma ◽  
Nabora Reyes de Mochel ◽  
Paula Duyen Pham ◽  
Tae Yeon Yoo ◽  
Ken W. Y. Cho ◽  
...  
Keyword(s):  
Fluorescence Lifetime ◽  
Embryo Quality ◽  
Assisted Reproductive Technologies ◽  
Learning Algorithm ◽  
Reproductive Technologies ◽  
Fluorescence Lifetime Imaging Microscopy ◽  
Fluorescence Lifetime Imaging ◽  
Embryo Viability ◽  
Experimental Conditions ◽  
Lifetime Imaging

Abstract Development of quantitative, safe and rapid techniques for assessing embryo quality provides significant advances in Assisted Reproductive Technologies (ART). Instead of assessing the embryo quality by the standard morphologic evaluation, we apply the phasor-FLIM (Fluorescence Lifetime Imaging Microscopy) method to capture endogenous fluorescent biomarkers of pre-implantation embryos as a non-morphological caliber for embryo quality. Here, we identify, under hypoxic and non-hypoxic conditions, the unique spectroscopic trajectories at different stages of mouse pre-implantation development, which is referred to as the developmental, or “D-trajectory”, that consists of fluorescence lifetime from different stages of mouse pre-implantation embryos. The D-trajectory correlates with intrinsic fluorescent species from a distinctive energy metabolism and oxidized lipids, as seen with Third Harmonic Generation (THG) that changes over time. In addition, we have defined a non-morphological Embryo Viability Index (EVI) to distinguish pre-implantation embryo quality using the Distance Analysis (DA), a machine learning algorithm to process the fluorescence lifetime distribution patterns. We show, under our experimental conditions, that the phasor-FLIM approach provides a much-needed non-invasive quantitative technology for identifying healthy embryos at the early compaction stage with 86% accuracy. The DA and phasor-FLIM method may provide the opportunity to improve implantation success rates for in vitro fertilization clinics.

scholarly journals In vivo measurement of pH and CO2 levels in the uterus of sows through the estrous cycle and after insemination

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Octavio López-Albors ◽  
Pedro José Llamas-López ◽  
Joaquín Ángel Ortuño ◽  
Rafael Latorre ◽  
Francisco Alberto García-Vázquez
Keyword(s):  
Estrous Cycle ◽  
Assisted Reproductive Technologies ◽  
Reproductive Technologies ◽  
Invasive Approach ◽  
Vitro Fertilization ◽  
Non Invasive ◽  
Physiological Values

AbstractThe pH–CO2–HCO3− system is a ubiquitous biological regulator with important functional implications for reproduction. Knowledge of the physiological values of its components is relevant for reproductive biology and the optimization of Assisted Reproductive Technologies (ARTs). However, in situ measurements of these parameters in the uterus are scarce or null. This study describes a non-invasive method for in situ time-lapse recording of pH and CO2 within the uterus of non-anesthetized sows. Animals were at three different reproductive conditions, estrous with no insemination and two hours after insemination, and diestrous. From pH and CO2 data, HCO3− concentration was estimated. The non-invasive approach to the porcine uterus with novel optical probes allowed the obtaining of in situ physiological values of pH, CO2, and HCO3−. Variable oscillatory patterns of pH, CO2 and HCO3− were found independently of the estrous condition. Insemination did not immediately change the levels of uterine pH, CO2 (%) and HCO3− concentration, but all the values were affected by the estrous cycle decreasing significantly at diestrous condition. This study contributes to a better understanding of the in vivo regulation of the pH-CO2-HCO3− system in the uterus and may help to optimize the protocols of sperm treatment for in vitro fertilization.

scholarly journals Metabolic state of human blastocysts measured by fluorescence lifetime imaging microscopy

2021 ◽  
Author(s):  
Marta Venturas ◽  
Jaimin S Shah ◽  
Xingbo Yang ◽  
Tim H Sanchez ◽  
William Conway ◽  
...  
Keyword(s):  
Fluorescence Lifetime ◽  
Cell Mass ◽  
Reproductive Technologies ◽  
Metabolic Imaging ◽  
Fluorescence Lifetime Imaging Microscopy ◽  
Fluorescence Lifetime Imaging ◽  
Embryo Viability ◽  
Metabolic State ◽  
Lifetime Imaging ◽  
Non Invasive

Mammalian embryos undergo large changes in metabolism over the course of preimplantation development. Embryo metabolism has long been linked to embryo viability, suggesting its potential utility in Assisted Reproductive Technologies (ART) to aid in selecting high quality embryos. However, the metabolism of human embryos remains poorly characterized due to a lack of non-invasive methods to measure their metabolic state. Here, we explore the application of metabolic imaging via fluorescence lifetime imaging microscopy (FLIM) for studying human blastocysts. We use FLIM to measure the autofluorescence of two central coenzymes, NAD(P)H and FAD+, in 215 discarded human blastocysts from 137 patients. We find that FLIM is sensitive enough to detect significant metabolic differences between blastocysts. We show that the metabolic state of human blastocysts changes continually over time, and that variations between blastocyst are partially explained by both the time since fertilization and their developmental stage, but not their morphological grade. We also observe significant metabolic heterogeneity within individual blastocysts, including between the inner cell mass and the trophectoderm, and between the portions of hatching blastocysts within and without the zona pellucida. Taken together, this work reveals novel aspects of the metabolism of human blastocysts and suggests that FLIM is a promising approach to assess embryo viability through non-invasive, quantitative measurements of their metabolism.

scholarly journals TERRA: A Novel Biomarker of Embryo Quality and Art Outcome

Genes ◽  
2021 ◽  
Vol 12 (4) ◽  
pp. 475
Author(s):  
Maria Santa Rocca ◽  
Ludovica Dusi ◽  
Andrea Di Nisio ◽  
Erminia Alviggi ◽  
Benedetta Iussig ◽  
...  
Keyword(s):  
Telomere Length ◽  
Dna Sequences ◽  
Male Fertility ◽  
Embryo Quality ◽  
Assisted Reproductive Technologies ◽  
Biological Clock ◽  
Reproductive Technologies ◽  
Age Related ◽  
Non Coding Rna ◽  
First Time

Telomeres are considered to be an internal biological clock, and their progressive shortening has been associated with the risk of age-related diseases and reproductive alterations. Over recent years, an increasing number of studies have focused on the association between telomere length and fertility, identifying sperm telomere length (STL) as a novel biomarker of male fertility. Although typically considered to be repeated DNA sequences, telomeres have recently been shown to also include a long non-coding RNA (lncRNA) known as TERRA (telomeric repeat-containing RNAs). Interestingly, males with idiopathic infertility show reduced testicular TERRA expression, suggesting a link between TERRA and male fertility. The aim of this study was to investigate the role of seminal TERRA expression in embryo quality. To this end, STL and TERRA expression were quantified by Real Time qPCR in the semen of 35 men who underwent assisted reproductive technologies (ART) and 30 fertile men. We found that TERRA expression in semen and STL was reduced in patients that underwent ART (both p < 0.001). Interestingly, TERRA and STL expressions were positively correlated (p = 0.010), and TERRA expression was positively associated with embryo quality (p < 0.001). These preliminary findings suggest a role for TERRA in the maintenance of sperm telomere integrity during gametogenesis, and for the first time, TERRA expression was found as a predictive factor for embryo quality in the setting of assisted reproduction.

scholarly journals Intrinsic and Extrinsic Factors Predicting the Cumulative Outcome of IVF / ICSI Treatment

2019 ◽  
Vol 9 (2S) ◽  
pp. 269-273 ◽  
Keyword(s):  
Assisted Reproductive Technologies ◽  
Sperm Quality ◽  
Reproductive Technologies ◽  
Process Time ◽  
Success Rates ◽  
Embryo Viability ◽  
Extrinsic Factors ◽  
Response Level ◽  
Female Population ◽  
Incident Cases

Infertility rates in India becoming increased in last decade principally due to the urbanization conditions and the lifestyle habits. It is giving alarm by continuously reporting the progress in incident cases of infertility amongst the young Indian adults of both male and female population. Among the various Assisted Reproductive Technologies (ART) available today in the treatment of infertility, In Vitro Fertilization (IVF) is found to be the most applicable treatment method of choice. This involves the administration of different hormones and drugs to treat infertility. In the present scenario technically IVF treatment process is tedious, laborious, high cost and most importantly success rates reported to be very low (20-30%). The prediction of IVF success rates is becoming an important scientific knowledge and practice, which helps both the doctor and the candidate couple to know about the conditions hence to take the right decision. The accurate prediction of the IVF success rate is really a challenging task in obstetrics and gynecology medicine. The success rates of the IVF depends on the various factors such as Intrinsic factors i.e, Genetic predisposition, Age, Body mass Index, Hormonal balance, Embryo viability, Sperm quality, Endometriosis and overall patient’s response level of the candidate couple and the Extrinsic factors such as Medical equipment technology, Treatment methods, Personal experiences of clinicians and embryologists, Process time, Stress due to the lifestyle etc.

1 Assessing the energy status of porcine embryos by means of biodynamic imaging

2020 ◽  
Vol 32 (2) ◽  
pp. 125
Author(s):  
I. Lorenzo ◽  
Z. Li ◽  
M. Torres ◽  
Z. Machaty ◽  
D. Nolte
Keyword(s):  
Sodium Azide ◽  
Assisted Reproductive Technologies ◽  
Imaging System ◽  
Reproductive Technologies ◽  
Optical Measurements ◽  
Energy Status ◽  
Embryo Viability ◽  
Cell Stage ◽  
Developmental Potential ◽  
Bioluminescence Assay

Assisted reproductive technologies are powerful tools for enhancing production in livestock or treating infertility in humans. Unfortunately, the success rate of the technologies is rather low. A major reason for the poor efficiency is the lack of methods to reliably assess the developmental potential of the embryos before transfer into recipients. Therefore, a noninvasive method to ensure the selection of only the best embryos for transfer would be highly desirable. Biodynamic imaging is a compelling new microscopy that uses intracellular Doppler spectroscopy to perform label-free, noninvasive optical measurements of cellular fitness. The aim of this study was to investigate whether biodynamic imaging can be used to assess the energy status of the embryos, which may be indicative of their viability. Porcine oocytes matured invitro were parthenogenetically activated by an electrical pulse and cultured for 2 days. The parthenotes were then divided into two groups, and approximately half of them were incubated for an additional 2 days in the presence of 20mM sodium azide. Sodium azide is an inhibitor of oxidative phosphorylation and is known to block ATP production. The rest of the embryos were cultured without sodium azide and used as a control to indicate normal ATP levels. At the end of the culture period embryos that reached the 8- to 16-cell stage were evaluated by our biodynamic imaging system to assess their energy status, after which they were lysed and their ATP contents were determined by means of a bioluminescence assay. A total of 68 embryos (32 treated with the inhibitor and 36 control) were evaluated. The ATP content analysis showed that the control embryos had significantly more ATP than those treated with sodium azide as determined by Student's t-test (5.04±1.07 vs. 1.31±0.57; P&lt;0.05). A correlative study was then completed where biodynamic biomarkers were used to classify embryos to estimate the ability of biodynamic imaging to identify embryos with high or low energy status. A set of 13 biomarkers representing each embryo as a feature vector was used to train a classifier. We found that the cross-validated classifier had a sensitivity and specificity of ~80%. In addition, a receiver-operator curve constructed by varying the ATP threshold of the independent bioluminescence assay had an area-under-the-curve of 0.81. These results indicate that biodynamic imaging is able to determine the energy status of the embryos noninvasively and has great potential in the assessment of embryo viability.

NON-INVASIVE METHODS FOR STUDYING THE POTENTIAL OF HUMAN EMBRYO FOR IMPLANTATION

2021 ◽  
pp. 29-37
Author(s):  
Василий Николаевич Попов ◽  
Роман Борисович Стукалин ◽  
Валерия Александровна Грибанова
Keyword(s):  
Assisted Reproductive Technologies ◽  
Culture Media ◽  
Uterine Cavity ◽  
Married Couples ◽  
Reproductive Technologies ◽  
Preimplantation Embryos ◽  
Preimplantation Genetic Testing ◽  
Vitro Fertilization ◽  
Non Invasive

В статье проводится анализ представленных на сегодня инвазивных и неинвазивных методов исследования преимплантационных эмбрионов. Показана эффективность преимплантационного генетического тестирования эмбрионов до переноса в полость матки. Также рассмотрены альтернативные менее инвазивные варианты изучения жизнеспособности эмбрионов, которые могли бы являться маркерами успешной имплантации. Проблема бесплодного брака с каждым годом становится все более и более значимой. Для части супружеских пар единственной возможностью рождения ребенка становится лечение методами вспомогательных репродуктивных технологий, эффективность которых остается на сегодняшний день не более 50 %. Особенно важным является поиск новых методик, позволяющих повысить результативность процедур экстракорпорального оплодотворения. В этом направлении крайне интересным является изучение неизвазивных методов оценки имплантационного потенциала эмбрионов. В анализе представлены работы по изучению протеома, метаболома и транскриптома эмбриона. Понимание молекулярного состава культуральных сред, в которых происходило развитие эмбриона до пятых суток культивирования, позволит глубже понять физиологию раннего развития, а также установить неивазивные критерии отбора эмбриона с лучшим имплантационным потенциалом и тем самым повысить эффективность проводимых программ вспомогательных репродуктивных технологий The article analyzes the currently presented invasive and non-invasive methods for studying preimplantation embryos. The efficiency of preimplantation genetic testing of embryos before transfer to the uterine cavity has been shown. Also considered are alternative less invasive options for studying the viability of embryos, which could be markers of successful implantation. The problem of sterile marriage is becoming more and more significant every year. For some married couples, the only possibility of having a child is treatment with methods of assisted reproductive technologies, the effectiveness of which remains at most 50% today. It is especially important to search for new techniques to improve the effectiveness of in vitro fertilization procedures. In this direction, it is extremely interesting to study non-invasive methods for assessing the implantation potential of embryos. The analysis presents works on the study of the proteome, metabolome and transcriptome of the embryo. Understanding the molecular composition of the culture media in which the development of the embryo took place until the fifth day of cultivation will allow a deeper understanding of the physiology of early development and also establish non-invasive criteria for the selection of embryos with the best implantation potential and thereby increase the efficiency of the programs of assisted reproductive technologies

scholarly journals Label-free microscopy: A non-invasive new tool to assess gametes and embryo quality

2020 ◽  
Vol 150 ◽  
pp. 241-246
Author(s):  
Marcello Rubessa ◽  
Matthew B. Wheeler
Keyword(s):  
Embryo Quality ◽  
Label Free ◽  
Non Invasive

scholarly journals Impact of myo-inositol treatment in women with polycystic ovary syndrome in assisted reproductive technologies

2021 ◽  
Vol 18 (1) ◽  
Author(s):  
Philippe Merviel ◽  
Pandora James ◽  
Sarah Bouée ◽  
Mathilde Le Guillou ◽  
Camille Rince ◽  
...  
Keyword(s):  
Polycystic Ovary Syndrome ◽  
Embryo Quality ◽  
Ovarian Function ◽  
Assisted Reproductive Technologies ◽  
Polycystic Ovary ◽  
Reproductive Technologies ◽  
Review Of The Literature ◽  
Ovary Syndrome ◽  
Randomized Controlled ◽  
The Impact

AbstractPolycystic ovary syndrome (PCOS) is marked in 30 to 40% by insulin resistance and hyperandrogenism. Myo-inositol (MI) increases insulin sensitivity, decreases hyperandrogenism and improves the menstrual cycle. Its effect during assisted reproductive technologies (ART) has been studied by many authors. We conducted a review of the literature on the impact of MI administration in PCOS women in assisted reproductive technologies. Myo-inositol is effective in normalizing ovarian function, improving oocyte and embryo quality in PCOS, however further evaluations by large multicentre randomized controlled trials are needed to assess the clinical pregnancy and live birth rates in ART.

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