![Figure 3. Increased placental oxidative stress in CBA/J × DBA/2 mice. Increased STAT-8 levels were observed in day-15 placentas from DBA/2-mated CBA/J mice compared with control matings (*P < .001). Treatment with hirudin (hir) or fondaparinox (FPX) prevented STAT-8 increase in CBA/J × DBA/2 mice (n = 6-8 mice/experimental group). Treatment with anti-TF 1H1 antibody or with Cl2MDP clodronate also prevented placental STAT-8 increase (n = 6-8 mice/experimental group). (B) Increased superoxide (O2−) production (DHE red staining) was observed in CBA/J × DBA/2 mice (i). O2− production was attenuated in CBA/J × DBA/2 mice treated with anti-TF (iii) or with Cl2MDP (iv). Minimal O2− production was observed in placentas from control matings (ii). (C) Macrophages from nonpregnant CBA/J female mice were stimulated with C5a, LPS, anti-TF, anti-TF + C5a, or control medium. Blockade of TF or genetic deletion of TF (TFfloxed/floxed/LysM-Cre mice) prevented C5a-induced release of sFlt-1 (n = 4-6 experiments/group; *P < .005 vs control). (D) Trophoblast proliferation assays. SM9-1 cells were incubated in RPMI-1640 as described before.21 Incubation with supernatants of monocytes incubated with C5a inhibited cell proliferation (ii). Incubation with increasing doses of sFlt-1 showed a dose-response inhibitory effect on SM9-1 cell proliferation ([iii] 2000 pg/mL, [iv] 4000 pg/mL, [v] 8000 pg/mL). Increased proliferation was observed when VEGF was added to the supernatant of C5a-treated monocytes (vi). (E) Superoxide (O2−) production in SM9-1 cells. Abundant SM9-1 cells with weak DHE-positive staining were observed in SM9-1 incubated with media (i). Fewer cells but strongly positive with DHE staining were observed in SM9-1 cells incubated with monocyte supernatants (ii) or sFlt-1 (4000 pg/mL [iii] and 8000 pg/mL [iv]). Decreased O2− production was observed when VEGF was added to the supernatant of C5a-treated monocytes (v). (F) Immunocytochemical analysis of TF. A large number of confluent SM9-1 cells with weak TF staining was observed after incubation with medium (i). Diminished cell proliferation and strong positive TF staining were observed in SM9-1 cells incubated with supernatants from C5a-treated monocytes (ii), sFlt-1 4000 pg/mL (iii), or sFlt-1 8000 pg/mL (iv). Increased proliferation and decreased TF were observed when VEGF was added to the supernatant of C5a-treated monocytes (v). Data are mean values plus or minus SD.](https://ash.silverchair-cdn.com/ash/content_public/journal/blood/113/17/10.1182_blood-2008-12-194258/4/zh80200934880003.jpeg?Expires=1723191875&Signature=sFuqkOZy47mLIMMfbyoimiVnSdx3m2KFjFyNqBcPKoBLKk6fKZRPaSPimiZuMoQMM8vCgsnG-PhEKyK4GML~5AcY~XfwUFCcEoJdnhUP6wHtZ1-DyIlHnxCwJrhc5kQtWh3TH598J3wrPwak5AWr0CY6d09o7YwDqpB-M6YkUMHfReN5GuCl4MOv4zlM~IN9uo0vjWjeVcLMhi6U0cax6Ns2DLjHly2Ej-SuT9jwZroGkG6dpwtZ5LZVeTnDewdULHMDTRx34zT5bv1WHAezto5XbKPh34ePVMAKxrB1fX0IB291tw8FQX9MXjPPNrETFXYbNSLakFTpgjYlV8Pg0g__&Key-Pair-Id=APKAIE5G5CRDK6RD3PGA)
Increased placental oxidative stress in CBA/J × DBA/2 mice. Increased STAT-8 levels were observed in day-15 placentas from DBA/2-mated CBA/J mice compared with control matings (*P < .001). Treatment with hirudin (hir) or fondaparinox (FPX) prevented STAT-8 increase in CBA/J × DBA/2 mice (n = 6-8 mice/experimental group). Treatment with anti-TF 1H1 antibody or with Cl2MDP clodronate also prevented placental STAT-8 increase (n = 6-8 mice/experimental group). (B) Increased superoxide (O2−) production (DHE red staining) was observed in CBA/J × DBA/2 mice (i). O2− production was attenuated in CBA/J × DBA/2 mice treated with anti-TF (iii) or with Cl2MDP (iv). Minimal O2− production was observed in placentas from control matings (ii). (C) Macrophages from nonpregnant CBA/J female mice were stimulated with C5a, LPS, anti-TF, anti-TF + C5a, or control medium. Blockade of TF or genetic deletion of TF (TFfloxed/floxed/LysM-Cre mice) prevented C5a-induced release of sFlt-1 (n = 4-6 experiments/group; *P < .005 vs control). (D) Trophoblast proliferation assays. SM9-1 cells were incubated in RPMI-1640 as described before.21 Incubation with supernatants of monocytes incubated with C5a inhibited cell proliferation (ii). Incubation with increasing doses of sFlt-1 showed a dose-response inhibitory effect on SM9-1 cell proliferation ([iii] 2000 pg/mL, [iv] 4000 pg/mL, [v] 8000 pg/mL). Increased proliferation was observed when VEGF was added to the supernatant of C5a-treated monocytes (vi). (E) Superoxide (O2−) production in SM9-1 cells. Abundant SM9-1 cells with weak DHE-positive staining were observed in SM9-1 incubated with media (i). Fewer cells but strongly positive with DHE staining were observed in SM9-1 cells incubated with monocyte supernatants (ii) or sFlt-1 (4000 pg/mL [iii] and 8000 pg/mL [iv]). Decreased O2− production was observed when VEGF was added to the supernatant of C5a-treated monocytes (v). (F) Immunocytochemical analysis of TF. A large number of confluent SM9-1 cells with weak TF staining was observed after incubation with medium (i). Diminished cell proliferation and strong positive TF staining were observed in SM9-1 cells incubated with supernatants from C5a-treated monocytes (ii), sFlt-1 4000 pg/mL (iii), or sFlt-1 8000 pg/mL (iv). Increased proliferation and decreased TF were observed when VEGF was added to the supernatant of C5a-treated monocytes (v). Data are mean values plus or minus SD.
