Expression of Regulatory T cell (Treg) Activation Markers in Endometrial Tissues from Early and Late Pregnancy in the Feline Immunodeficiency Virus (FIV)-Infected Cat

TaqMan real time RT-PCR analysis of FIV gag gene expression in placentas and corresponding fetuses from early (week 3–4) pregnancy and placentas from late (week 8) pregnancy in FIV-infected cats. (a) Viral mRNA was amplified from all placental samples (14 of 14 samples). FIV gag was detected in 12 of 14 fetal samples. The asterisks indicate that no fetal tissue was collected from placenta 5111 C, 8035 D, and 1893 R due to resorption. (b) Viral RNA was detected in 10 of 17 placental samples from the late term FIV-infected queens. Negative placental samples 6108 A, 9276 A, 9522 A, and 9746 A were obtained from uninoculated, control queens. Bars (adjusted mean Ct) represent mean Ct values subtracted from a negative endpoint (50).

TaqMan real time RT-PCR analysis of endometrial expression of Treg markers CD25, FOXP3, and CTLA4 in early gestation control (n = 18) versus late gestation control (n = 13) samples. Bars (adjusted mean Ct) represent mean Ct values subtracted from a negative endpoint (50), and error bars represent standard errors of the mean. P values < 0.05 were considered significant.

TaqMan real time RT-PCR analysis of expression of Treg markers CD25, FOXP3, and CTLA4 in control and infected reproductive tissues at early and late pregnancy. Samples were evaluated as follows: infected (n = 17) versus control (n = 18) at early pregnancy; and infected (n = 18) versus control (n = 13) at late pregnancy. Bars (adjusted mean Ct) represent mean Ct values subtracted from a negative endpoint (50), bracketed by standard errors of the mean. P values < 0.05 were considered significant.

Relative expression of CD25, FOXP3, and CTLA4 in early and late term infected reproductive tissues producing viable versus non-viable fetuses. The samples were evaluated as follows: infected cats producing viable offspring at early pregnancy (n = 11) versus infected cats producing non-viable offspring at early pregnancy (n = 5); and infected cats producing viable offspring at late pregnancy (n = 9) versus infected cats producing non-viable offspring at late pregnancy (n = 9). Bars (adjusted mean Ct) represent mean Ct values subtracted from a negative endpoint (50), bracketed by standard errors of the mean. P values < 0.05 were considered significant.

Immunofluorescence labeling of FOXP3 at the maternal–fetal interface of tissue from a representative control queen. FOXP3 (green) was detected using polyclonal rabbit anti-FOXP3 antiserum followed by goat anti-rabbit IgG (H + L) fluorescein conjugate. Cells were counterstained with DAPI (blue). Negative controls included universal negative control antibody or no primary antibody, resulting in no fluorescent labeling (data not shown). Cells were viewed by confocal laser scanning microscopy using a 40× oil immersion objective.