Impact of extracellular folic acid levels on oviductal gene expression

MANSILLA, MARIANO J.; GARCÍA, ELINA V.; BARRERA, ANTONIO D. ; LEBLANC, JEAN G.

THERIOGENOLOGY

ELSEVIER SCIENCE INC

Lugar: Amsterdam; Año: 2020 vol. 154 p. 161 - 170

0093-691X

Folate plays a specific role as methyl donor for nucleotidesynthesis and genomic methylation patterns, which in turn are importantepigenetic determinants in gene expression. Previous studies haverevealed the presence of folate in bovine oviductal fluid as well as theexistence of a fine-tuned regulation of the gene expression of folatereceptors and transporters in bovine oviduct epithelial cells (BOECs).However, the functional implications of folate in the oviduct remainunknown. The present study aimed to assess the effect of folic acid (FA)on expression levels of selected genes that potentially respond to thefolate status in in vitro BOECs. To obtain an insight into theoptimization of a culture system for assays, gene expression of folatereceptors and transporters was compared between BOECs grown in monolayersand in suspension. The results showed that BOECs from isthmus and ampullain suspension culture better preserved the region-dependent geneexpression profile than in monolayers. Subsequently, BOECs from bothanatomical regions were separately cultured in suspension for 24 hassaying different FA concentrations: I) TCM-199 (control); II) TCM-199 +1 μM FA (similar to the oviduct concentration); III) TCM-199 + 10 μM FAand IV) TCM-199 + 100 μM FA. Expression analysis of genes related toimportant cellular processes including folate transport, DNA methylation,cell-cell interaction, antioxidant activity and signaling pathways wasperformed in BOECs using RT-qPCR. Our data demonstrated that addition of1 μM FA did not affect mRNA levels of most genes analyzed. In contrast,BOECs cultured with 10 μM FA exhibited increased mRNA expression levelsof genes involved in folate intake, DNA methylation and antioxidantprotection. It is worth noting that at 100 μM FA, transcriptionalresponse in BOECs mainly resulted in decreased mRNA levels of themajority of the genes assayed. Interestingly, cytotoxicity analysisshowed a similar LDH activity in the culture media of the experimentalgroups, indicating that cell integrity was not affected by the FAconcentrations assayed. In conclusion, our findings suggest that folatecan affect BOECs, promoting changes in gene activity in a framework offunctional readjustments in response to environmental conditions.