DOI: 10.6092/unibo/amsacta/7210
title: ConcePTION WP3 task 3.2. Isolation and characterization of Mammary Epithelial Cells from Göttingen minipig (mpMECs): an in vitro study to compare mpMECs vs pMECs (porcine Mammary Epithelial Cells)

description: This data set contains data related to the characterization of an appropriate in vitro animal model based on primary culture of Göttingen Minipig Mammary Epithelial Cells (mpMECs). The objective of the research was to verify the accuracy of mpMECs as solid translational model for the study of mammary epithelial barrier and compare mpMECs results with those obtained from Mammary Epithelial Cells isolated from hybrid commercial pig (pMECs). The results showed that it was possible to isolate, culture and expand three pure epithelial cell lines obtained from three different animals. The mpMECs maintained until P10 showed a typical cobblestone morphology and a similar doubling time profile (MG9 32.6 ± 4 h, MG11 30.5 ± 3 h and MG12 27.4 ± 2.8 h, as reported in the data file CONCEPTION_WP3_mpMECs_DoublingTime_09032023.xlsx). DNA index was normal for all the three cell populations with a mean value of 0.98 ± 0.01. Regarding the cell cycle, the cell populations MG9, MG11 and MG12 showed the three distinct phases that could be recognized in a proliferating cell population: G0/G1, S and G2/M (see data file CONCEPTION_WP3_mpMECs_CellCycle_DNAIndex_09032023.xlsx). All the three mpMECs cell lines expressed epithelial markers Epithelial-Cadherin (E-Cad) and Cytokeratin 18 (CK18), confirming their epithelial origin (see data file CONCEPTION_WP3_mpMECs_FlowCytometer_E-Cad_CK18_09032023.xlsx). In particular in MG12, the contour of CK18 positive peak showed a shoulder suggesting the presence of a cellular subpopulation with a particularly high positivity. The barrier function of mpMECs was evaluated via TEER and fluorescein sodium (SF) transport, the formation of the monolayer integrity was evaluated in all the three primary mpMECs and in the pre-mixed pool of mpMECs and pMECs. MG9, MG11 and MG12 resulted in a similar TEER profile at the 0.15×10^6 cells, achieving higher TEER and lower SF values at the day 3 and 4 of culture, which was also confirmed in the pre-mixed pool case. Moreover, by comparing the maximum values of TEER reached at the different seeding density tested, a significative difference resulted in MG9 with respect to MG11, MG12 and pool. From a similar comparison with minimum values of SF transport, no difference between the groups was showed. Finally, in pre-mixed pools of mpMECs and pMECs a difference in the ability to form the epithelial barrier was shown: mpMEC barrier resulted less tight compact then that formed by pMECs. (see data file CONCEPTION_WP3_mpMECs_TEER_SF_09032023.xlsx). On pre-mixed pool of mpMECs and pMECs we evaluated the bioenergetic metabolism: no difference in energy production under basal cell culture condition was observed but under stressed state pMECs made more efficient use of mitochondrial oxidative metabolism than mpMECs, conversely, these last ones could more efficiently increased the glycolytic activity (see data file CONCEPTION_WP3_mpMECs_BioenergerticMetabolism_09032023.xlsx). The transcriptional profile of drug transporters evaluated in pre-mixed pool of mpMECs or pMECs cultured MECGM medium. Among the 84 genes, 66 genes were detectable, 18 genes were not detectable or higher than 35 threshold cycle, so considered as negative according to the handbook, in both cell lines. No difference between mpMECs and pMECs drug transporter gene expression levels was observed (see data file CONCEPTION_WP3_mpMECs_RT2ARRAY_09032023.xlsx).