A microfluidic in vitro cultivation system for mechanical stimulation of bovine embryos

Abstract

This work demonstrates a novel microfluidic in vitro cultivation system for embryos that improves their development using a partially constricted channel that mimics peristaltic muscle contraction. Conventional photolithography and a PDMS replica molding process were used to make straight or constricted microchannels. To investigate the effects of constriction geometry on embryonic development, different constriction widths of the channel were designed. Bovine embryos were loaded and incubated by simply placing them on a tilting machine to provide embryo movement via gravity. The fertilized embryos were cultivated on the microfluidic in vitro cultivation system until the blastocyst, hatching, or hatched blastocyst stages. To confirm the quality of blastocysts in the microfluidic channel, double staining was performed and compared with bovine embryos cultivated by the conventional droplet method. The proportion of eight-cell development among total embryos in the constricted channel (56.7+/-13.7%; mean+/-SD) was superior to that in the straight channel (23.9+/-11.0%). This suggests that the effect of constriction is vital for the early development of bovine embryos in assisted-reproduction research.

Culture of Bovine Embryos on a Polydimethylsiloxane (PDMS) Microwell Plate

“…PDMS has been used in biological studies, not only because of its easy fabrication and lowcost, but also because it is impermeable to water, nontoxic to cells and permeable to gases [24]. Recently, Kim et al [25] showed that the quality of bovine IVF embryos cultured in a PDMS-based microfluidic in vitro cultivation system was comparable with those cultured using a conventional method. The results of this past study [25] and those of the present study suggest that culture on PDMS has no harmful effects on the developmental ability and quality of bovine embryos.…”

“…Recently, Kim et al [25] showed that the quality of bovine IVF embryos cultured in a PDMS-based microfluidic in vitro cultivation system was comparable with those cultured using a conventional method. The results of this past study [25] and those of the present study suggest that culture on PDMS has no harmful effects on the developmental ability and quality of bovine embryos. However, it is necessary to examine whether embryos cultured on a PDMS-MP support full development to term after embryo transfer.…”

An integrated microfluidic array system for evaluating toxicity and teratogenicity of drugs on embryonic zebrafish developmental dynamics

“…7,8 To date, a number of biomedical application, such as cell-based investigation, in particular, cell cultivation and proliferation has been shown to great success relied on microfluidic chip. 9,10 Apart from cells, [11][12][13] embryos, [14][15][16][17][18] nematodes, [19][20][21][22] and tissues, 23,24 all have realized on-chip cultivation or manipulation. However, the microscale channels fail to easily handle the multicellular organism of large size, such as zebrafish embryos, which might obscure their promise of application in compound-related toxicity and teratogenicity assay.…”

Use of Silicone Elastomer-Based Microfluidic Devices and Systems in Reproductive Technologies

“…Within the last decade, studies using PDMS microfluidic channels or funnels have suggested novel solutions for oocyte manipulation, sperm sorting, and embryo culture . Microfluidic systems that mimic oviductal structures and functions for use in RT are divided into those used for sperm motility control or monitoring [50][51][52][53][54][55][56][57][58][59][60], regulation of chemical gradients for in vitro fertilization and embryo culture [61][62][63][64][65][66][67][68][69], and applying mechanical stimuli to the developing embryo [70][71][72][73][74]. Although the unique characteristics of elastomers have been exploited for microstructure fabrication and/or micro-pumping, mechanical deformation of the elastomer membrane has not been used to produce new devices for RT.…”

“…Progression to the blastocyst developmental stage was significantly enhanced under dynamic microfunnel culture conditions, as evidenced by an increased percentage of hatching or hatched blastocysts and a significantly higher average number of cells per blastocyst. Kim et al developed a DCS using a combination of a PDMS microfluidic channel and a tilting machine, as shown in Figure 6C [72]. Bovine embryos were loaded and incubated by simply placing them on a tilting machine to provide embryo movement via gravity.…”