Toll-like receptor 9 activation: a novel mechanism linking placenta-derived mitochondrial DNA and vascular dysfunction in pre-eclampsia

Abstract

Emerging evidence suggests that in addition to being the 'power houses' of our cells, mitochondria facilitate effector responses of the immune system. Cell death and injury result in the release of mtDNA (mitochondrial DNA) that acts via TLR9 (Toll-like receptor 9), a pattern recognition receptor of the immune system which detects bacterial and viral DNA but not vertebrate DNA. The ability of mtDNA to activate TLR9 in a similar fashion to bacterial DNA stems from evolutionarily conserved similarities between bacteria and mitochondria. mtDNA may be the trigger of systemic inflammation in pathologies associated with abnormal cell death. PE (pre-eclampsia) is a hypertensive disorder of pregnancy with devastating maternal and fetal consequences. The aetiology of PE is unknown and removal of the placenta is the only effective cure. Placentas from women with PE show exaggerated necrosis of trophoblast cells, and circulating levels of mtDNA are higher in pregnancies with PE. Accordingly, we propose the hypothesis that exaggerated necrosis of trophoblast cells results in the release of mtDNA, which stimulates TLR9 to mount an immune response and to produce systemic maternal inflammation and vascular dysfunction that lead to hypertension and IUGR (intra-uterine growth restriction). The proposed hypothesis implicates mtDNA in the development of PE via activation of the immune system and may have important preventative and therapeutic implications, because circulating mtDNA may be potential markers of early detection of PE, and anti-TLR9 treatments may be promising in the management of the disease.

Figure 1

A synthetic TLR9 ligand (ODN 2395; InvivoGen, San Diego, CA, USA) was administered by intraperitoneal injection (0.1 µg/rat) on days 14, 17, and 19 of gestation (term=21 days) in pregnant rats or corresponding days in non-pregnant rats. Blood pressure was measured via the tail cuff method on gestational day 20 and rats were euthanized on day 21. A) Systolic blood pressure of non-pregnant (NP) and late pregnant rats (Preg) treated with ODN 2395 (NP-ODN 2395, n=1; Preg-ODN 2395, n=2) or Vehicle (NP-Veh, n=3; Preg-Veh, n=2), B) Fetal weights from pregnant rats treated with ODN 2395 or Vehicle.

Figure 2

Mitochondria were isolated from rat liver (Mitochondria Isolation Kit, Pierce Biotechnology, Rockford, IL, USA) and their integrity was disrupted by sonication. Mitochondria solution (4 mg tissue/rat diluted in saline) was injected in pregnant rats on gestational day 15. Blood pressure was measured via the tail cuff method on gestational day 18 and rats were euthanized on day 19. A) Systolic blood pressure of pregnant rats injected with mitochondria (Preg-mt, n=2) or Vehicle (Preg-Veh, n=2), B) Densitometric intensity and representative Western blots for TLR9 protein) in relation to β-actin in second order mesenteric arteries from pregnant rats injected with mitochondria (Preg-mt, n=2) or Vehicle (Preg-Veh, n=2), C) Densitometric intensity and representative Western blots for phosphorylated ERK1/2 proteins in relation to total ERK1/2 in mesenteric arteries from pregnant rats injected with mitochondria (Preg-mt, n=2) or Vehicle (Preg-Veh, n=2).

Figure 3

Diagram demonstrating the hypothesis that during pregnancy placenta-derived mitochondrial DNA, through TLR9 activation, increases vascular reactivity to constrictor stimuli via an ERK1/2-dependent signaling pathway and potentiates the release of pro-inflammatory cytokines, contributing to preeclampsia-like symptoms (i.e., maternal hypertension and intrauterine growth restriction).