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Pyrroloquinoline quinone (PQQ) supplementation during in vitro culture of murine embryos alters mitochondrial activity but has minimal effects on embryo development

      Objective

      Pyrroloquinoline quinone (PQQ) has been proposed as a culture medium supplement to support embryo mitochondrial function and mitigate the negative effects of reactive oxygen species (ROS) in vitro, particularly in embryos of women of advanced maternal age. The aim of this study was to determine whether PQQ supplementation during mouse embryo culture is beneficial, and how PQQ may affect embryo mitochondrial function.

      Design

      Research study.

      Materials and Methods

      First, a high (2.5 uM) and low (0.25 uM) concentration of PQQ during the culture period (112h) was investigated. Two-cell embryos were assessed for mitochondrial activity using JC-1. Remaining embryos were cultured to the blastocyst stage and analyzed for cell number and allocation, and ATP; this was common across experiments. Next, 0.25 uM PQQ was used in step 1, step 2, or throughout a sequential culture system. Two-cell embryos were removed for ROS staining. Finally, oocytes were collected from aged (15 mo) and young (1 mo) mice and cultured in 0 uM or 0.25 uM PQQ in step one of a sequential culture system. Two-cell embryos were stained with JC-1. Significance was determined at p<0.05.

      Results

      There were no differences in blastocyst development or cell allocation at either level of PQQ supplementation. There were also no differences in ATP production, which was the case in all experiments. An increase was observed in mitochondrial activity (n=38) of 2-cell embryos following culture in 0.25 uM PQQ. There was no difference in blastocyst development depending on when or if PQQ was supplemented, or in abundance of ROS. Blastocyst total cell number was increased when embryos were cultured with PQQ in step one only compared to control (132.7 ± 5.2 and 114.4 ± 6.7 respectively; n=74). More embryos from aged females hatched when PQQ was included in culture step 1 (n=301). Embryos from aged mice cultured with PQQ had increased mitochondrial activity compared to those without PQQ (n=40).

      Conclusions

      Supplementation of embryo culture medium with PQQ did not result in production of more blastocysts, though blastocysts resulting from culture with PQQ in step 1 only had more total cells. In addition, more blastocysts from aged females hatched when PQQ was included in culture step 1. PQQ consistently increased mitochondrial activity in two-cell embryos. These results suggest that improvement in mitochondrial metabolism in early cleavage stages may provide some benefit to blastocyst quality.