Anti-Oxidative Effects of Human Adipose Stem Cell Conditioned Medium with Different Basal Medium during Mouse Embryo In Vitro Culture

Abstract

The quality of embryos produced by assisted reproductive techniques should be advanced by the improvement of in vitro culture conditions for successful implantation and pregnancy maintenance. We investigated the anti-oxidative effect of human adipose stem cell (ASC) conditioned medium with its optimal basal medium, Dulbecco′s modified Eagle′s medium (DMEM-CM), or keratinocyte serum-free medium (KSFM-CM) as supplements during in vitro culture (IVC) of in vitro fertilized mouse embryo. At first, preimplantation embryo development was evaluated in KSFM-CM and DMEM-CM supplemented cultures at various concentrations. The blastocyst (BL) and hatched BL formation rates were significantly increased in 5% DMEM-CM, while no difference was observed from KSFM-CM. Next, comparing the efficacy of KSFM-CM and DMEM-CM at the same concentration, DMEM-CM enhanced the developmental rate of 16 cells, morula, BL, and hatched BL. The expression level of reactive oxygen species decreased and that of glutathione increased in BL cultured with DMEM-CM, which confirms its anti-oxidative effect. Furthermore, apoptosis in BL cultured with DMEM-CM was reduced compared with that in KSFM-CM. This study demonstrated that the comparative effect of human ASC-CM made of two different basal media during mouse embryo IVC and anti-oxidative effect of 5% DMEM-CM was optimal to improve preimplantation embryo development.

Comparison of Anti-Oxidative Effect of Human Adipose- and Amniotic Membrane-Derived Mesenchymal Stem Cell Conditioned Medium on Mouse Preimplantation Embryo Development

“…The rate of embryos that developed to the 4-cell and 16-cell stage were significantly lower in the 50% AMSC-CM group than in the 10% AMSC-CM group. These results indicate that a high CM concentration does not ensure better efficiency of embryo development regardless of the large quantity of cytokines as previously explained [6]. Remarkably, we found that 10% AMSC-CM significantly improved BL formation rate (Table 2), which is an index for embryo developmental potential and consequently determines the success of implantation [41], compared to the control group as well as the other higher concentration of AMSC-CM treated groups.…”

“…2019-03). ASCs were cultured and characterized as previously described [ 6 ]. For the establishment of AMSCs, cryopreserved AMSCs (1 × 10 6 ) from the amnion tissue of three female donors were cultured in T-175 flasks containing RPME-P (R BIO, Seoul, Korea) supplemented with 1% antibiotic-antimycotic solution at 37 °C with 5% CO 2 .…”

“…ASC-CM were collected using the same method as previously described [ 6 ]. AMSCs per donor (passage 6) were cultured in RPME-P until 90% confluency, and then the medium was replaced to Dulbecco’s modified Eagle’s medium (DMEM) after washing twice with PBS.…”

“…In the process of assisted reproduction, a number of external factors causing oxidative stress appear from technical procedures to environmental sources [ 3 ]. Subsequently, oxidative stress due to accumulated ROS in in vitro-produced embryos impairs the efficiency of embryonic development and induces reproductive failure due to an increase in embryo fragmentation and apoptosis, and a decrease in fertilization rate and blastocyst (BL) development [ 4 , 5 , 6 ]. Accordingly, the application of antioxidants to ART can be an effective intervention to counteract the oxidative damage in in vitro-produced embryos [ 7 ], especially to improve the in vitro culture (IVC) medium for favorable outcomes in preimplantation embryo development.…”

“…MSCs isolated from multiple tissue sources have common but various features, which emphasize their importance as regenerative medicine [ 14 ]. Among multiple sources, adipose-derived MSCs (ASCs) are the most widely studied, forming the basis of research on MSC-based therapy [ 15 ] and is representatively known a strong antioxidant [ 6 , 13 , 16 , 17 , 18 ]. Contemporarily, amniotic membrane-derived MSCs (AMSC) have emerged as a novel candidate in the field of regenerative medicine because of their unique advantages, including noninvasive isolation, stable immunogenicity, abundant availability, multipotency for all three germ layers, and no associated ethical issues [ 19 ].…”