RUPP rats

RUPP rats. Effect of sFlt-1/PlGF on vascular and uteroplacental collagen types I and IV. banding the lower abdominal aorta above the iliac bifurcation and main uterine branches of the ovarian arteries as previously explained (16, 29, 101). Briefly, the abdominal aorta near the iliac bifurcation was dissected free of perivascular excess fat and separated from your vena cava. A blunt plastic rod (outer diameter: 0.3 mm) was placed parallel to the aorta, and a 4-0 silk braided ligature was knotted twice round the aorta and adjacent plastic rod. Once taut, the rod was removed from the knotted ligature, thus creating a constrictive band (inner diamter: 0.3 mm) and reducing blood flow through the aorta. This procedure reduces uterine perfusion pressure in the gravid rat by 40% (19). Because compensation of blood flow to the placenta occurs through adaptive increase in ovarian blood flow (62), a blunt plastic rod (outer diameter: 0.1 mm) was used to place a ligature band (inner diameter: 0.1 mm) on the main uterine branches of both the right and left ovarian arteries, specifically avoiding the main ovarian artery. RUPP rats received postoperative analgesia in the form of subcutaneous buprenorphine (0.05 mg/kg) every 12 h for 48 h plus meloxicam (2 mg/kg) every 24 h for 48 h. RUPP rats in which the banding process resulted in maternal death or total reabsorption of the pups were excluded from data analyses, and the RUPP process success rate was ~85%. Normal Preg rats were sham operated. Some RUPP rats were simultaneously infused intravenously via a jugular vein catheter and osmotic pump with recombinant PlGF-1 (MBS696135, MyBiosource, San Diego, CA) at 20 gkg?1day?1 for 5 days (RUPP + PlGF) (101). A lower dose of PlGF (10 gkg?1day?1) was not sufficient to decrease BP in RUPP rats. A previous study infused recombinant VEGF at 90 or 180 gkg?1day?1 to restore the angiogenic sense of balance and showed that it lowered BP and improved renal function in RUPP rats (30). We selected PlGF over VEGF because most studies have shown decreased PlGF levels in PE (8, 53, 89), whereas measurements of VEGF have not been consistent, with studies showing decreased (68), no switch (54, 55), or even increased levels (9, 34, 89). Also, PlGF is usually specific for VEGFR-1 and its soluble form sFlt-1, whereas VEGF also binds to VEGFR-2 and could increase vascular permeability and edema (49) and promote malignancy (77). VEGF levels are also controlled at the maternal-fetal Isoliquiritigenin interface, partly through opinions modulation of sFlt-1, to prevent damage to the placenta or fetus by extra VEGF (21), and dysregulation of this feedback mechanism could complicate the measurement of angiogenic factors. A recent study also showed that infusion of recombinant human PlGF at 180 gkg?1day?1 abolished placental ischemia-induced HTN-Preg in rats (87). We avoided using higher doses of PlGF, as they may cause microvascular abnormalities (39). Also, because the angiogenic/antiangiogenic balance is usually tightly controlled by opinions mechanisms, Isoliquiritigenin extra PlGF could drive a feedback increase in sFlt-1 to maintain the sFlt-1-to-PlGF ratio. We used a smaller dose of 20 gkg?1day?1 PlGF based on our guiding measurements of sFlt-1 and PlGF levels in Preg, RUPP, and Preg + sFlt-1 rats, and the dose of PlGF was sufficient to counterbalance sFlt-1 and decrease the sFlt-1-to-PlGF ratio in RUPP + PlGF versus RUPP rats to levels close to those in control Preg rats. All procedures followed the National Institutes of Health and the guidelines of the American Physiological Society and were approved by the Institutional Animal Care and Use Committee of the Brigham and Women Hospital. Blood pressure. On of pregnancy, rats were anaesthetized with isoflurane, and a PE-50 catheter was inserted in the carotid artery and exteriorized at the back of the neck. Rats were allowed to recover from anesthesia for at least 1 h. Isoliquiritigenin The carotid arterial catheter was connected to a pressure transducer attached to an amplifier and pressure recorder (Living System Instrumentation, Burlington, VT). Animals were not tethered and did not receive analgesia during measurement of KLF1 BP. BP in conscious rats was monitored over a 30-min period, and the average BP was measured as previously explained (56). Plasma sFlt-1 and PlGF. After BP had been measured, blood samples were collected via the arterial catheter into sterile heparin tubes (Tyco Healthcare, Mansfield, MA), and plasma was separated by centrifugation at 2,000?for 10 min and stored at ?80C. Plasma sFlt-1 levels were measured using rat sFlt-1 ELISA microplate kit (MBS2602003, MyBiosource), with 0.05 ng/ml sFlt-1 sensitivity, intra-assay and interassay precision, and a coefficient.