bims-gamemb Biomed News
on Gamete and embryo metabolism
Issue of 2021–12–26
eight papers selected by
Cameron A. Schmidt, East Carolina University
  1. Inhibition of DRP1 Impedes Zygotic Genome Activation and Preimplantation Development in Mice.
  2. Sperm-Guiding Unconventional Prostaglandins in C. elegans: Synthesis and Signaling.
  3. MFN1 and MFN2 Are Dispensable for Sperm Development and Functions in Mice.
  4. Concurrent Measurement of Mitochondrial DNA Copy Number and ATP Concentration in Single Bovine Oocytes.
  5. Design of novel oocyte activation methods: The role of zinc.
  6. Hypoxic in vitro culture reduces histone lactylation and impairs pre-implantation embryonic development in mice.
  7. Effects of different stimulators of cGMP synthesis on lipid content in bovine oocytes matured in vitro.
  8. Inherited Metabolic Memory of High-fat Diet Impairs Testicular Fatty Acid Content and Sperm Parameters.
  1. Front Cell Dev Biol. 2021 ;9 788512
    Inhibition of DRP1 Impedes Zygotic Genome Activation and Preimplantation Development in Mice.
    Yuanyuan Li, Ning-Hua Mei, Gui-Ping Cheng, Jing Yang, Li-Quan Zhou.
      Mitochondrion plays an indispensable role during preimplantation embryo development. Dynamic-related protein 1 (DRP1) is critical for mitochondrial fission and controls oocyte maturation. However, its role in preimplantation embryo development is still lacking. In this study, we demonstrate that inhibition of DRP1 activity by mitochondrial division inhibitor-1, a small molecule reported to specifically inhibit DRP1 activity, can cause severe developmental arrest of preimplantation embryos in a dose-dependent manner in mice. Meanwhile, DRP1 inhibition resulted in mitochondrial dysfunction including decreased mitochondrial activity, loss of mitochondrial membrane potential, reduced mitochondrial copy number and inadequate ATP by disrupting both expression and activity of DRP1 and mitochondrial complex assembly, leading to excessive ROS production, severe DNA damage and cell cycle arrest at 2-cell embryo stage. Furthermore, reduced transcriptional and translational activity and altered histone modifications in DRP1-inhibited embryos contributed to impeded zygotic genome activation, which prevented early embryos from efficient development beyond 2-cell embryo stage. These results show that DRP1 inhibition has potential cytotoxic effects on mammalian reproduction, and DRP1 inhibitor should be used with caution when it is applied to treat diseases. Additionally, this study improves our understanding of the crosstalk between mitochondrial metabolism and zygotic genome activation.
    Keywords:  DRP1; ROS; mitochondrial dysfunction; preimplantation development; zygotic genome activation
    DOI:  https://doi.org/10.3389/fcell.2021.788512
  2. Metabolites. 2021 Dec 08. pii: 853. [Epub ahead of print]11(12):
    Sperm-Guiding Unconventional Prostaglandins in C. elegans: Synthesis and Signaling.
    Ekta Tiwary, Muhan Hu, Jeevan K Prasain.
      Prostaglandins comprise a family of lipid signaling molecules derived from polyunsaturated fatty acids and are involved in a wide array of biological processes, including fertilization. Prostaglandin-endoperoxide synthase (a.k.a. cyclooxygenase or Cox) initiates prostaglandin synthesis from 20-carbon polyunsaturated fatty acids, such as arachidonic acid. Oocytes of Caenorhabditis elegans (C. elegans) have been shown to secrete sperm-guidance cues prostaglandins, independent of Cox enzymes. Both prostaglandin synthesis and signal transduction in C. elegans are environmentally modulated pathways that regulate sperm guidance to the fertilization site. Environmental factors such as food triggers insulin and TGF-β secretion and their levels regulate tissue-specific prostaglandin synthesis in C. elegans. This novel PG pathway is abundant in mouse and human ovarian follicular fluid, where their functions, mechanism of synthesis and pathways remain to be established. Given the importance of prostaglandins in reproductive processes, a better understanding of how diets and other environmental factors influence their synthesis and function may lead to new strategies towards improving fertility in mammals.
    Keywords:  C. elegans; Cox-independent pathway; TGF-β; prostaglandin; sperm guidance
    DOI:  https://doi.org/10.3390/metabo11120853
  3. Int J Mol Sci. 2021 Dec 16. pii: 13507. [Epub ahead of print]22(24):
    MFN1 and MFN2 Are Dispensable for Sperm Development and Functions in Mice.
    Junru Miao, Wei Chen, Pengxiang Wang, Xin Zhang, Lei Wang, Shuai Wang, Yuan Wang.
      MFN1 (Mitofusin 1) and MFN2 (Mitofusin 2) are GTPases essential for mitochondrial fusion. Published studies revealed crucial roles of both Mitofusins during embryonic development. Despite the unique mitochondrial organization in sperm flagella, the biological requirement in sperm development and functions remain undefined. Here, using sperm-specific Cre drivers, we show that either Mfn1 or Mfn2 knockout in haploid germ cells does not affect male fertility. The Mfn1 and Mfn2 double knockout mice were further analyzed. We found no differences in testis morphology and weight between Mfn-deficient mice and their wild-type littermate controls. Spermatogenesis was normal in Mfn double knockout mice, in which properly developed TRA98+ germ cells, SYCP3+ spermatocytes, and TNP1+ spermatids/spermatozoa were detected in seminiferous tubules, indicating that sperm formation was not disrupted upon MFN deficiency. Collectively, our findings reveal that both MFN1 and MFN2 are dispensable for sperm development and functions in mice.
    Keywords:  MFN1; MFN2; haploid spermatids; male fertility; sperm functions
    DOI:  https://doi.org/10.3390/ijms222413507
  4. Methods Protoc. 2021 Dec 07. pii: 88. [Epub ahead of print]4(4):
    Concurrent Measurement of Mitochondrial DNA Copy Number and ATP Concentration in Single Bovine Oocytes.
    Casey C Read, Sadikshya Bhandari, Sarah E Moorey.
      To sustain energy-demanding developmental processes, oocytes must accumulate adequate stores of metabolic substrates and mitochondrial numbers prior to the initiation of maturation. In the past, researchers have utilized pooled samples to study oocyte metabolism, and studies that related multiple metabolic outcomes in single oocytes, such as ATP concentration and mitochondrial DNA copy number, were not possible. Such scenarios decreased sensitivity to intraoocyte metabolic relationships and made it difficult to obtain adequate sample numbers during studies with limited oocyte availability. Therefore, we developed and validated procedures to measure both mitochondrial DNA (mtDNA) copy number and ATP quantity in single oocytes. Validation of our procedures revealed that we could successfully divide oocyte lysates into quarters and measure consistent results from each of the aliquots for both ATP and mtDNA copy number. Coefficient of variation between the values retrieved for mtDNA copy number and ATP quantity quadruplicates were 4.72 ± 0.98 and 1.61 ± 1.19, respectively. We then utilized our methodology to concurrently measure mtDNA copy number and ATP quantity in germinal vesicle (GV) and metaphase two (MII) stage oocytes. Our methods revealed a significant increase in ATP levels (GV = 628.02 ± 199.53 pg, MII = 1326.24 ± 199.86 pg, p < 0.001) and mtDNA copy number (GV = 490,799.4 ± 544,745.9 copies, MII = 1,087,126.9 ± 902,202.8 copies, p = 0.035) in MII compared to GV stage oocytes. This finding is consistent with published literature and provides further validation of the accuracy of our methods. The ability to produce consistent readings and expected results from aliquots of the lysate from a single oocyte reveals the sensitivity and feasibility of using this method.
    Keywords:  ATP; mitochondria; mitochondrial DNA; oocyte; single cell analysis
    DOI:  https://doi.org/10.3390/mps4040088
  5. Biol Reprod. 2021 Dec 22. pii: ioab235. [Epub ahead of print]
    Design of novel oocyte activation methods: The role of zinc.
    Kyungjun Uh, Alayna Hay, Paula Chen, Emily Reese, Kiho Lee.
      Oocyte activation occurs at the time of fertilization and is a series of cellular events initiated by intracellular Ca2+ increases. Consequently, oocytes are alleviated from their arrested state in meiotic metaphase II (MII), allowing for the completion of meiosis. Oocyte activation is also an essential step for somatic cell nuclear transfer (SCNT) and an important tool to overcome clinical infertility. Traditional artificial activation methods aim to mimic the intracellular Ca2+ changes which occur during fertilization. Recent studies emphasize the importance of cytoplasmic Zn2+ on oocyte maturation and the completion of meiosis, thus suggesting artificial oocyte activation approaches that are centered around the concentration of available Zn2+in oocytes. Depletion of intracellular Zn2+ in oocytes with heavy metal chelators leads to successful oocyte activation in the absence of cellular Ca2+ changes, indicating that successful oocyte activation does not always depends on intracellular Ca2+ increases. Current findings lead to new approaches to artificially activate mammalian oocytes by reducing available Zn2+ contents, and the approaches improve the outcome of oocyte activation when combined with existing Ca2+ based oocyte activation methods. Here, we review the important role of Ca2+ and Zn2+ in mammalian oocyte activation and development of novel oocyte activation approaches based on Zn2+ availability.
    Keywords:  Oocyte activation; Zinc; embryo development; fertilization
    DOI:  https://doi.org/10.1093/biolre/ioab235
  6. Epigenetics Chromatin. 2021 Dec 21. 14(1): 57
    Hypoxic in vitro culture reduces histone lactylation and impairs pre-implantation embryonic development in mice.
    Wanting Yang, Peijun Wang, Pengbo Cao, Shuang Wang, Yuxiao Yang, Huimin Su, Buhe Nashun.
       BACKGROUND: Dynamic changes of histone posttranslational modifications are important contexts of epigenetic reprograming after fertilization in pre-implantation embryos. Recently, lactylation has been reported as a novel epigenetic modification that regulates various cellular processes, but its role during early embryogenesis has not been elucidated.
    RESULTS: We examined nuclear accumulation of H3K23la, H3K18la and pan histone lactylation in mouse oocytes and pre-implantation embryos by immunofluorescence with specific antibodies. All of the three modifications were abundant in GV stage oocytes, and both H3K23la and pan histone lactylation could be detected on the condensed chromosomes of the MII oocytes, while H3K18la were not detected. After fertilization, the nuclear staining of H3K23la, H3K18la and pan histone lactylation was faint in zygotes but homogeneously stained both of the parental pronuclei. The signal remained weak in the early cleavage stage embryos and increased remarkably in the blastocyst stage embryos. Comparison of the embryos cultured in four different conditions with varying concentrations of oxygen found that H3K23la, H3K18la and pan histone lactylation showed similar and comparable staining pattern in embryos cultured in atmospheric oxygen concentration (20% O2), gradient oxygen concentration (5% O2 to 2% O2) and embryos obtained from in vivo, but the modifications were greatly reduced in embryos cultured in hypoxic condition (2% O2). In contrast, nuclear accumulation of H3K18ac or H3K23ac was not significantly affected under hypoxic condition. Moreover, the developmental rate of in vitro cultured embryo was significantly reduced by low oxygen concentration and small molecule inhibition of LDHA activity led to decreased lactate production, as well as reduced histone lactylation and compromised developmental rate.
    CONCLUSIONS: We provided for the first time the dynamic landscape of H3K23la, H3K18la and pan histone lactylation in oocytes and pre-implantation embryos in mice. Our data suggested that histone lactylation is subjected to oxygen concentration in the culture environment and hypoxic in vitro culture reduces histone lactylation, which in turn compromises developmental potential of pre-implantation embryos in mice.
    Keywords:  Embryonic development; H3K18la; H3K23la; Histone lactylation; Hypoxia; Oxygen; Pre-implantation embryo
    DOI:  https://doi.org/10.1186/s13072-021-00431-6
  7. Anim Reprod. 2021 ;18(4): e20210072
    Effects of different stimulators of cGMP synthesis on lipid content in bovine oocytes matured in vitro.
    Letícia Schefer, Kátia Regina Lancelloti Schwarz, Daniela Martins Paschoal, Fernanda Cavallari de Castro, Hugo Fernandes, Ramon César Botigelli, Cláudia Lima Verde Leal.
      Bovine oocytes and blastocysts produced in vitro are frequently of lower quality and less cryotolerant than those produced in vivo, and greater accumulation of lipids in the cytoplasm has been pointed out as one of the reasons. In human adipocytes cGMP signaling through the activation of PKG appears to be involved in lipid metabolism, and components of this pathway have been detected in bovine cumulus-oocyte complexes (COCs). The aim of this study was to investigate the influence of this pathway on the lipid content in oocytes and expression of PLIN2 (a lipid metabolism-related gene) in cumulus cells. COCs were matured in vitro for 24 h with different stimulators of cGMP synthesis. The activation of soluble guanylyl cyclase (sGC) by Protoporphyrin IX reduced lipid content (22.7 FI) compared to control oocytes (36.45 FI; P <0.05). Stimulation of membrane guanylyl cyclase (mGC) with natriuretic peptides precursors A and C (NPPA and NPPC) had no effect (36.5 FI; P>0.05). When the PKG inhibitor KT5823 was associated with Protoporphyrin IX, its effect was reversed and lipid contents increased (52.71 FI; P<0.05). None of the stimulators of cGMP synthesis affected the expression of PLIN2 in cumulus cells. In conclusion, stimulation of sGC for cGMP synthesis promotes lipolytic activities in bovine oocytes matured in vitro and such effect is mediated by PKG. However, such effect may vary depending on the stimulus received and/or which synthesis enzyme was activated, as stimulation of mGC had no effects.
    Keywords:  NPR1; Nile red; guanylate cyclase; lipid droplets; natriuretic peptides
    DOI:  https://doi.org/10.1590/1984-3143-AR2021-0072
  8. Mol Nutr Food Res. 2021 Dec 23. e2100680
    Inherited Metabolic Memory of High-fat Diet Impairs Testicular Fatty Acid Content and Sperm Parameters.
    Luís Crisóstomo, Romeu A Videira, Ivana Jarak, Kristina Starčević, Tomislav Mašek, Luís Rato, João F Raposo, Rachel L Batterham, Pedro F Oliveira, Marco G Alves.
      Exposure to a high-fat diet (HFD) from early-life is associated with a testicular metabolic signature linked to abnormal sperm parameters up to two generations after exposure in mice. Hereby, we describe a testicular lipid signature associated with "inherited metabolic memory" of exposure to HFD, persisting up to two generations in mice. Diet-challenged mice (n = 36) were randomly fed after weaning with standard chow (CTRL); HFD for 200 days or transient HFD (HFDt ) (60 days of HFD+140 days of standard chow). Subsequent generations (36 mice per generation) were fed with chow diet. Mice were euthanized 200 days post-weaning. Glucose homeostasis, serum hormones, testicular bioenergetics and antioxidant enzyme activity were evaluated. Testicular lipid-related metabolites and fatty acids were characterized by 1 H-NMR and GC-MS. Sons of HFD display impaired choline metabolism, mitochondrial activity and antioxidant defenses, while grandsons show a shift in testicular ω3/ω6 ratio towards a pro-inflammatory environment. Grandsons of HFDt raise 3-hydroxybutyrate levels with possible implications to testicular insulin resistance. Sperm counts decrease in grandsons of HFD-exposed mice, regardless of the duration of exposure. HFD-induced "inherited metabolic memory" alters testicular fatty acid metabolism with consequences to sperm parameters up to two generations. This article is protected by copyright. All rights reserved.
    Keywords:  discriminant analysis; high-fat diet; inherited metabolic memory; lipidomics; paternal inheritance; testis
    DOI:  https://doi.org/10.1002/mnfr.202100680
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