Inflammatory immune response of canine trophoblastic cells from primary culture to infection with Brucella canis

FERNANDEZ, AG; HIELPOS, MS; FERRERO, MC; FOSSATI, CA; BALDI, PC

Mar del Plata

Simposio; V Latin American Symposium on Reproductive Immunology (LASRI); 2015

Upto date, Brucella canis is the only Brucella specie which has been related tocanine abortion. It has also been documented the transmission to human beings.However, the pathogenic mechanisms by which this coccobacillus causes abortionhaven´t been described so far.Objectives: 1) to confirmB. canis ability to infect andreplicate inside canine trophoblastic cells from primary culture; 2) to analyzethe inflammatory immune response triggered by infectionMethods: canine trophoblastic cellswere isolated from term placenta obtained by C-section of healthy female dogs andcharacterized by immune fluorescence with anti-cytokeratine 7 (ck7) antibody.5x104 cells/well were infected with B. canis M+ at MOI 250 for 48hs. After treatment withgentamicin,cells were lysed and plated onto bacteriological agar for CFUdetermination at 2, 24 and 48h post infection (p.i.). Culture supernatants werecollected at 24 and 48h p.i. Cytotoxicity was evaluated through LDH release andcytokines were measured by ELISA.Results: Isolated canine trophoblastswere positive for ck7 staining. Replication kinetic showed a significantdecrease (p<0.05) in CFU/well between 2h and 24h p.i., followed by anotorious increase (p<0.01) at 48h p.i. No differences in LDH levels weredetected between uninfected and infected cells. Following infection, both IL-8and RANTES secretion was significantly higher at 48h p.i (p<0.05 andp<0.01, respectively), while IL-6 secretion remained undetectable.Conclusions: canine trophoblastsinfected with B. canis generate aninflammatory immune response, mediated by IL-8 and RANTES, which may contributeto uterine local damage leading to fetal miscarriage.