Deletion of Junctional Adhesion Molecule a (JAM-A) Renders Protection to Mice from Severe Sepsis in a Sex Dependent Manner

Introduction: Sepsis is the body's extreme response to an infection which leads to multiple organ dysfunction and mortality. Males are more susceptible to sepsis than females. It is believed that sex hormones may be responsible for this difference. Recent studies highlight the role of platelets in sepsis. Thrombocytopenia and thrombosis are the major events that occur during sepsis. We have shown that JAM-A, a cell adhesion molecule belonging to Ig superfamily is expressed in platelets and regulates agonist-induced platelet activation by suppressing α _IIbβ ₃ activation in mice. JAM-A is also expressed on leucocytes and endothelial cells, where it regulates leukocyte extravasation and vascular permeability. However, the role of JAM-A in sepsis is poorly understood.

Aim: To evaluate the role of Jam-A in polymicrobial sepsis using a murine model of sepsis.

Methods: Global Jam-A knock out( Jam-A ^gt/gt) mice were generated using genetrap. Severe sepsis was induced in Jam-A ^gt/gt mice endothelial specific Jam-A ^{fl/fl/Tie2Cre}, andcorresponding control (WT or Jam-A ^fl/fl) mice by cecal ligation (100%) and puncture (CLP) model. Bacterial colony forming units (CFU) in blood and peritoneal lavage were enumerated using blood agar plates. VEGF levels were assessed by ELISA.

Results: We observed that upon CLP both male and female WT and male Jam-A ^gt/gt (Jam-A null) mice showed high septic shock score and severe hypothermia. On the contrary, the female Jam-A ^gt/gtmice showed lower septic shock score and no sign of hypothermia. Within 48 h of CLP 100% WT mice (both male and female n=10) died. Similarly, 100% Jam-A ^gt/gt male mice (n=10) also died within 48 h of CLP. Surprisingly, 100% Jam-A ^gt/gt female mice (n=10) survived until the end of the study (7 days). These results suggest that Jam-A suppresses the protective advantage of female sex. When evaluated for bacteremia by enumerating CFU in blood we found the presence of numerous bacteria in the blood of all WT mice and male Jam-A ^gt/gt mice within 24 h of CLP. Interestingly, none of the female Jam-A ^gt/gt mice had any bacterial presence in the blood at 24 h time suggesting that bacterial entry through the endothelium may be restricted due to the loss of Jam-A in conjunction with female sex hormone signaling. Furthermore, the bacterial count was also significantly reduced in the peritoneum of female Jam-A ^gt/gt as opposed to WT and male Jam-A ^gt/gt mice suggesting that the bacteria were rapidly cleared in the peritoneum of female Jam-A ^gt/gt mice. Since, Jam-A is known to regulate vascular permeability and leukocyte transmigration, we performed CLP in mice lacking endothelial specific Jam-A ( Jam-A ^{fl/fl/Tie2Cre}) and Jam-A ^fl/flmice as control. We found that just like in WT mice, 100% male and female Jam-A ^fl/fl, and 100% male Jam-A ^{fl/fl/Tie2Cre} mice died within 48 h. In contrast, 100% female Jam-A ^{fl/fl/Tie2Cre} mice survived until the end of the study suggesting that endothelial Jam-A is responsible for subduing the protective function of female hormones. Since VEGF is the major factor regulating vascular permeability and plasma VEGF levels are increased in septic patients, we evaluated the plasma VEGF levels in mice after 24 h of CLP. As compared to male and female WT mice as well as Jam-A ^gt/gt male mice, an increased VEGF levels were observed in Jam-A ^gt/gt female mice. To evaluate if the increased VEGF levels were responsible for the rapid clearance of bacteria in the peritoneum due to efficient leukocyte transmigration, we pretreated WT mice with DC101, an anti-VEGFR2 antibody or isotype specific IgG (200μg/kg). We found that DC101 protected both male and female mice from severe sepsis and these mice had no bacteria in the blood as well as had no sign of septic shock suggesting that although VEGF supports bacteremia, the suppressive role of Jam-A in female mice is not due to VEGF.

Conclusion: Endothelial Jam-A plays an important role in regulating sepsis in female mice. Jam-A may suppress the protective function of female sex hormones. Although, VEGF positively contribute to bacteremia by enhancing endothelial permeability, the role of Jam-A in suppressing sex hormone function is independent of VEGF in mice.