Porcine Monocyte-Derived Macrophage Generation

Six cross-bred pigs (Sus scrofa domesticus) of either sex, aged 6–18 months old, were used as blood donors for in vitro experiments. Pigs were housed at the Experimental Station of Istituto Zooprofilattico Sperimentale (IZS) of Sardinia (“Surigheddu”, Sassari, Italy). Animal husbandry, handling, and procedures (bleeding) were carried out according to the Italian Legislative Decree No. 26 dated 4 March 2014 and in agreement with the Guide of Use of Laboratory Animals issued by the Italian Ministry of Health (authorization No. 1232/2020-PR).
Heparinized blood samples were used for generation of monocyte-derived macrophages (moMΦ) (described in Section 4.2). Animal health was routinely monitored by trained veterinarians, and blood samples were screened for several porcine pathogens. The absence of African swine fever (ASFV), porcine parvovirus (PPV), and porcine circovirus 2 (PCV2) genome was evaluated though qualitative real-time PCR, as previously described [21 (link),60 (link)], with primers reported in the Table S1 [61 (link),62 (link),63 (link)]. The absence of the porcine reproductive and respiratory syndrome virus (PRRSV) and Mycoplasma hyopneumoniae was monitored using commercial real-time PCR kits (LSI VetMAX™ PRRSV EU/NA and VetMAX™-Plus qPCR Master Mix, both Thermo Fisher Scientific, respectively), following the manufacturer’s instructions [21 (link)].