Analyzing mitochondrial respiration of human induced pluripotent stem cell-derived myeloid progenitors using Seahorse technology

Fan, Yanxin; Mizoguchi, Yoko; Tatematsu, Megumi; Linder, Monika I.; Frenz, Stephanie; Choi, Jong-Su; Klein, Christoph

doi:10.1016/j.xpro.2023.102073

Cited by 2 publications

(1 citation statement)

References 19 publications

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“…As sperm after cryopreservation are very sensitive to further handling within in vitro conditions both methods perfectly fit to this concept as they are live-cell, real-time, minimally invasive techniques [19]. The XF Analyzer is routinely used in the field of cancer [20], immunology [21], and stem cell research [22]. This method is important for sperm quality assessment, particularly in cases such as testicular cancer.…”

Section: Introductionmentioning

confidence: 99%

Bioenergetics of human spermatozoa in patients with testicular germ cell tumour

Simonik,

Bryndova,

Sur

et al. 2024

Preprint

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In testicular germ cell tumour (TGCT) patients, sperm cryopreservation prior to anti-cancer treatment represents the main fertility preservation approach. However, it is associated with low sperm recovery rate after thawing. Since sperm is a high-energy demanding cell, which is supplied by glycolysis and oxidative phosphorylation (OXPHOS), mitochondrial dysfunctionality can directly result in sperm anomalies. In this study, we investigated the bioenergetic pattern of cryopreserved sperm of TGCT patients in comparison with normozoospermic samples using two state-of-the-art methods; the Extracellular Flux Analyzer (XF Analyzer) and Two-Photon Fluorescence Lifetime imaging (2P-FLIM), in order to assess the contributions of OXPHOS and glycolysis to energy provision. A novel combined protocol for combined measurement of OXPHOS (Oxygen Consumption Rate – OCR) and glycolysis (Extracellular Acidification Rate – ECAR) using the XF Analyzer was developed together with a unique customized AI-based approach for semiautomated processing of 2P-FLIM images. Our study delivers optimized Low-HEPES modified Human Tubal Fluid media (mHTF) for sperm handling during pre-analytical and analytical phases to maintain sperm physiological parameters and optimal OCR, equivalent of OXPHOS. The negative effect of cryopreservation was signified by deterioration of both bioenergetic pathways represented by modified OCR and ECAR curves and the derived parameters. This was true for normozoospermic as well as TGCT samples, which showed even a stronger damage within the respiratory chain compared to the level of glycolytic activity impairment. These data are supported by 2P-FLIM analysis showing a significantly decreased bound NADH in contrast to unbound NAD(P)H which reflects decreased metabolic activity in samples from TGCT patients. Our study provides novel insight into the impact of TGCT on sperm bioenergetics and delivers a verified protocol to be used for assessment of human sperm metabolic activity, which can be a valuable tool for further research and clinical andrology.

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Section: Introductionmentioning

confidence: 99%

Bioenergetics of human spermatozoa in patients with testicular germ cell tumour

Simonik,

Bryndova,

Sur

et al. 2024

Preprint

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Generation of complex bone marrow organoids from human induced pluripotent stem cells

Frenz-Wiessner,

Fairley,

Buser

et al. 2024

Nat Methods

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The human bone marrow (BM) niche sustains hematopoiesis throughout life. We present a method for generating complex BM-like organoids (BMOs) from human induced pluripotent stem cells (iPSCs). BMOs consist of key cell types that self-organize into spatially defined three-dimensional structures mimicking cellular, structural and molecular characteristics of the hematopoietic microenvironment. Functional properties of BMOs include the presence of an in vivo-like vascular network, the presence of multipotent mesenchymal stem/progenitor cells, the support of neutrophil differentiation and responsiveness to inflammatory stimuli. Single-cell RNA sequencing revealed a heterocellular composition including the presence of a hematopoietic stem/progenitor (HSPC) cluster expressing genes of fetal HSCs. BMO-derived HSPCs also exhibited lymphoid potential and a subset demonstrated transient engraftment potential upon xenotransplantation in mice. We show that the BMOs could enable the modeling of hematopoietic developmental aspects and inborn errors of hematopoiesis, as shown for human VPS45 deficiency. Thus, iPSC-derived BMOs serve as a physiologically relevant in vitro model of the human BM microenvironment to study hematopoietic development and BM diseases.

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Analyzing mitochondrial respiration of human induced pluripotent stem cell-derived myeloid progenitors using Seahorse technology

Cited by 2 publications

References 19 publications

Bioenergetics of human spermatozoa in patients with testicular germ cell tumour

Bioenergetics of human spermatozoa in patients with testicular germ cell tumour

Generation of complex bone marrow organoids from human induced pluripotent stem cells

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