Mitochondrial energy metabolism is downregulated in repeated implantation failure patients related to alteration of PGC-1α acetylation level.

Herein we aimed at exploring mitochondrial energy metabolism status in patients with repeated implantation failure (RIF) and whether key regulatory factor PGC-1α of energy metabolism is involved in the decidualization of endometrial stromal cells. Mitochondrial oxidative phosphorylation level and ATP synthesis were compared in primary endometrial stromal cells from RIF and control group. At the same time, as one of key transcription regulators of mitochondrial energy metabolism, the expression level and acetylation level of PGC-1α were compared with two groups. Then, we downregulated the acetylation levels of PGC-1α, and the expression of decidual markers (PRL and IGFBP1) was observed further. Mitochondrial energy metabolism, showing by mitochondrial oxidative phosphorylation level and ATP synthesis, was decreased in the endometrial stromal cells of the RIF group (RIF-hEnSCs). Meanwhile, PGC-1α acetylation levels were significantly higher in RIF-hEnSCs. When we reduced the acetylation levels of PGC-1α in RIF-hEnSCs, the basal O2 consumption rate and maximal respiration were increased, and also the PRL and IGFBP1. Overall, our data indicated that the endometrial stromal cells of the RIF patients had low level of mitochondrial energy metabolism. Reducing acetylation level of key energy metabolism regulator PGC-1α can increase the decidualization level of RIF-hEnSCs. These findings may inspire new ideas about the treatment of RIF.

Effect of luteinizing hormone concentration on transcriptome and subcellular organelle phenotype of ovarian granulosa cells.

RESEARCH QUESTION: Granulosa cells (GCs) surrounding oocytes are crucial for follicular growth, oocyte development, ovulation, and luteinization under the dynamic co-stimulation of follicle stimulating hormone (FSH) and luteinizing hormone (LH). This study aimed to investigate the effect of LH levels on GCs in preovulatory follicles under gonadotropin releasing hormone antagonist-based ovarian stimulation. In vitro experiments were also conducted to study the direct effect of LH on GCs. METHODS: Twelve infertile women were divided into low (L), medium (M), and high (H) LH groups according to their serum LH levels during ovarian stimulation. RNA-sequencing (RNA-seq) was conducted to examine the transcriptome profiles of GCs obtained from the above patients during the oocyte retrieval. The activity of mitochondrial dehydrogenase was measured under the stimulation of recombinant LH (rLH) concentration gradient combined with recombinant FSH. The ultrastructures of subcellular organelles were observed. RESULTS: Bioinformatic analyses showed that compared with the M group, molecule and pathway changes in the L group and in the H group were similar. In cultured GCs, both insufficient and excessive rLH impaired the activity of mitochondrial dehydrogenase. With the medium rLH concentration, numerous cell connections and abundant mitochondria and liposomes were observed. Compared with the medium concentration, GCs showed smaller and rounder mitochondria, more autophagosomes, and massive organelles damages with excessive rLH, and swollen, circular, or forked mitochondria were observed with inadequate rLH. CONCLUSIONS: RNA-seq provided a novel spectrum of transcriptome characteristics of GCs potentially affected by serum LH levels during ovarian stimulation. In vitro, rLH could directly affect GCs at the subcellular level.

A modified holding pipette for mouse oocyte fertilization.

The safety of assisted reproductive technology (ART) is of frequent concern. Unfortunately, animal models for studying the safety of intracytoplasmic sperm injection (ICSI) have limitations in mimicking human ICSI manipulations. As reported herein, we invented a modified holding pipette for mouse oocyte injection that resulted in the delivery of live pups. A modified holding pipette was prepared for mouse oocyte injection and was compared with the conventional pipette for human use and a trumpet-shaped pipette. After ICSI, the oocytes were cultured to cleavage embryos until fallopian transfer. The use of the trumpet-shaped holding pipette and the new modified holding pipette for mouse oocyte injection achieved comparable and satisfactory oocyte survival rates (83.44% and 85.71%, respectively) and embryo cleavage rates (41.98% and 42.42%, respectively), which were significantly higher than those obtained with the human egg-holding pipette (oocyte survival rate: 65.85%; embryo cleavage rate: 27.78%). After 13 embryos were transferred using each type of pipette, three live pups were produced with the new modified holding pipette, one was produced with the holding pipette for human use, and none were produced with the trumpet-shaped holding pipette. The modified holding pipette for oocyte injection is effective and very easy to prepare. Moreover, using this new method, we produced live pups, which will contribute to a useful animal model for safety studies of ICSI in the future.

A novel Drosophila mitochondrial carrier protein acts as a Mg(2+) exporter in fine-tuning mitochondrial Mg(2+) homeostasis.

The homeostasis of magnesium (Mg(2+)), an abundant divalent cation indispensable for many biological processes including mitochondrial functions, is underexplored. In yeast, the mitochondrial Mg(2+) homeostasis is accurately controlled through the combined effects of importers, Mrs2 and Lpe10, and an exporter, Mme1. However, little is known about this Mg(2+) homeostatic process in multicellular organisms. Here, we identified the first mitochondrial Mg(2+) transporter in Drosophila, the orthologue of yeast Mme1, dMme1, by homologous comparison and functional complementation. dMme1 can mediate the exportation of mitochondrial Mg(2+) when heterologously expressed in yeast. Altering the expression of dMme1, although only resulting in about a 10% change in mitochondrial Mg(2+) levels in either direction, led to a significant survival reduction in Drosophila. Furthermore, the reduced survival resulting from dMme1 expression changes could be completely rescued by feeding the dMME1-RNAi flies Mg(2+)-restricted food or the dMME1-over-expressing flies the Mg(2+)-supplemented diet. Our studies therefore identified the first Drosophila mitochondrial Mg(2+) exporter, which is involved in the precise control of mitochondrial Mg(2+) homeostasis to ensure an optimal state for survival.

A novel mitochondrial carrier protein Mme1 acts as a yeast mitochondrial magnesium exporter.

The homeostasis of magnesium (Mg2+), an abundant divalent cation indispensable for many biological processes including mitochondrial functions, is underexplored. Previously, two mitochondrial Mg2+ importers, Mrs2 and Lpe10, were characterized for mitochondrial Mg2+ uptake. We now show that mitochondrial Mg2+ homeostasis is accurately controlled through the combined effects of previously known importers and a novel exporter, Mme1 (mitochondrial magnesium exporter 1). Mme1 belongs to the mitochondrial carrier family and was isolated for its mutation that is able to suppress the mrs2Δ respiration defect. Deletion of MME1 significantly increased steady-state mitochondrial Mg2+ concentration, while overexpression decreased it. Measurements of Mg2+ exit from proteoliposomes reconstituted with purified Mme1 provided definite evidence for Mme1 as an Mg2+ exporter. Our studies identified, for the first time, a mitochondrial Mg2+ exporter that works together with mitochondrial importers to ensure the precise control of mitochondrial Mg2+ homeostasis.

Mitochondrial release of the NADH dehydrogenase Ndi1 induces apoptosis in yeast.

Saccharomyces cerevisiae NDI1 codes for the internal mitochondrial ubiquinone oxidoreductase, which transfers electrons from NADH to ubiquinone in the respiratory chain. Previously we found that Ndi1 is a yeast homologue of the protein apoptosis-inducing factor-homologous mitochondrion-associated inducer of death and displays potent proapoptotic activity. Here we show that S. cerevisiae NDI1 is involved in apoptosis induced by various stimuli tested, including H(2)O(2), Mn, and acetate acid, independent of Z-VAD-fmk (a caspase inhibitor) inhibition. Although Ndi1 also participates in respiration, its proapoptotic property is separable from the ubiquinone oxidoreductase activity. During apoptosis, the N-terminal of Ndi1 is cleaved off in the mitochondria, and this activated form then escapes out to execute its apoptotic function. The N-terminal cleavage appears to be essential for the manifestation of the full apoptotic activity, as the uncleaved form of Ndi1 exhibits much less growth-inhibitory activity. Our results thus indicate an important role of Ndi1 in the switch of life and death fates in yeast: during normal growth, Ndi1 assimilates electrons to the electron transport chain and initiates the respiration process to make ATP, whereas under stresses, it cleaves the toxicity-sequestering N-terminal cap, is released from the mitochondria, and becomes a cell killer.