CD200Fc attenuates inflammatory responses and maintains barrier function by suppressing NF-κB pathway in cigarette...

Recent evidence suggests that CD200 fusion protein (CD200Fc), a CD200R1 agonist may attenuate inflammatory responses in autoimmune diseases and neuro-degeneration. While, little is known about the function of CD200Fc in cigarette smoke extract (CSE)-induced mouse Cardiac Microvascular Endothelial Cells (mCMECs). The present study was designed to elucidate the effects of CD200Fc on CSE-induced vascular endothelial barrier (VEB) dysfunction and inflammatory responses, which is a highly clinically relevant model of smoking related cardiovascular diseases.

mCMECs were pre-treated with 1, 10 and 100μg/ml CD200Fc for 24h respectively, and then treated with 250μg/ml CSE for different times (24h or 120min). The transepithelial electrical resistance (TEER) and transport of fluorescent markers were used to measure VEB function in CSE-induced mCMECs. Western blot and immunofluorescent staining analysis were used to detect the expression of tight junction proteins, such as Zona Occludens-1 (ZO-1) and Claudin-1 in CSE-induced mCMECs. We measured the expression of pro-inflammatory cytokines in CSE-induced mCMECs by using ELISA and RT-PCR. In addition, the NF-κB activity in CSE-induced mCMECs were investigated by using nuclear/cytosol fractionation and western blot analysis.

In vitro treatment with CSE increased the transport of fluorescent markers and decreased TEER levels in mCMECs, respectively, which were attenuated by CD200Fc (10 and 100μg/ml) pretreatment. The CSE-induced up-regulation of pro-inflammatory cytokines such as Cyclooxygenase-2 (COX-2), inducible nitric oxide synthase (iNOS), platelet endothelial cell adhesion molecule-1 (PECAM-1), vascular cell adhesion molecule-1 (ICAM-1), Prostaglandin E2 (PGE2), tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6) and IL-8 in mCMECs was also abrogated by CD200Fc (10 and 100μg/ml) pretreatment. CD200Fc also inhibited CSE-induced nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) activation in mCMECs, such as inhibition of its DNA binding activity, phosphorylated expression, and translocation to nucleus.

Thus, CD200Fc exert anti-inflammatory effect and protect VEB function in CSE-induced mCMECs. The vasoprotective effects of CD200Fc may be specifically beneficial in pathophysiological conditions associated with smoking related cardiovascular diseases.