Bovine peripheral blood WC1 + and WC1 neg γδ T lymphocytes modulate monocyte-derived macrophage effector functions during in vitro Mycobacterium avium subspecies paratuberculosis infection

Bovine WC1+ and WC1neg γδ T Lymphocytes Influence Monocyte Differentiation and Monocyte-Derived Dendritic Cell Maturation during In Vitro Mycobacterium avium Subspecies paratuberculosis Infection

“…In our system, MHC-II was not upregulated on mMDDCs at 48 h post Map infection; however, the expression of MHC-II was highly variable between animals in our study. Inter-animal immune cell phenotypic variability has been well established in bovine studies of our lab and others (25, 26, 49), and the distinct pattern of high/low-effector functions and specific phenotypes in cattle is complex but probably explains an individual’s unique ability to respond (or not) to infection. Timing may also have influenced our ability to detect early and transient changes in MHC-II expression.…”

“…Viability of Map recovered from dMonWC1 neg , however, was significantly reduced suggesting that the presence of WC1 neg γδ T lymphocytes improves the ability of differentiated monocytes (dMonWC1 neg ) to limit Map viability. In a previous study, we showed that the presence of either WC1 + or WC1 neg γδ T lymphocytes cocultured with autologous Map -infected MDMs from 30- to 40-day-old calves was associated with reduced viability of Map recovered from MDMs (26). Taking into account that (1) in the previous study, MDMs were considered fully differentiated prior to the addition of γδ T lymphocytes into cocultures, (2) in the current study, significant differences were observed between the viability of Map recovered from dMonWC1 + and dMonWC1 neg , and (3) when Map was introduced in the current study, WC1 + and WC1 neg γδ T lymphocytes had been removed; we hypothesize that dMonWC1 neg are distinct from classical MDMs, and that WC1 neg γδ T lymphocytes alter the functional differentiation of peripheral blood monocytes.…”

“…PBMCs were isolated from whole blood using Histopaque 1077 (Sigma Aldrich, Oakville, ON, Canada) density gradient centrifugation, counted using a Moxi Z cell counter (Orflo, Hailey, ID, USA), and resuspended in complete medium RPMI 1640 containing 2 mM of l -glutamine and 25 mM of HEPES (Gibco, Carlsbad, CA, USA) supplemented with 5 × 10 −5  M 2-mercaptoethanol (Sigma Aldrich, Oakville, ON, Canada), with penicillin (1,000 U/mL), streptomycin sulfate (10 mg/mL), and amphotericin B (0.25 µg/mL) (Sigma Aldrich). Our source of serum was 10% autologous serum based on our previous findings that show that cells are not self-reactive to cytokines or other soluble mediators present in autologous serum (26). Cell suspensions were transferred to 175 cm 2 flasks (Corning, Tewksbury, MA, USA) at a concentration of 7.5 × 10 6 /mL per flask.…”

“…During adaptive immune responses, γδ T lymphocytes develop memory responses (15, 16), induce DC maturation (17), and polarize into T H 1-, T H 2-, T H 17-, T FH -, or T REG -effector functions based on the cytokine milieu in which γδ T lymphocytes encounter the antigen (17–20). In cattle, γδ T lymphocytes have shown to produce pro-inflammatory cytokines, such as IFN-γ and IL-17A (18–21), regulate granuloma development (22), have regulatory effects (23, 24), and modulate macrophage-effector functions (25, 26). …”

“…Furthermore, we have previously shown that γδ T lymphocytes influence autologous monocyte-derived macrophage (MDM) effector functions of young calves and heifers during Map infection in vitro (25, 26). Therefore, the hypothesis for this study was that WC1 + and WC1 neg γδ T lymphocytes of young calves influence (1) monocyte differentiation and (2) DC maturation during Map infection in vitro .…”