Superoxide dismutase sod

For blood parameter measurements, GTT and ITT were tested as described above (6-h fast); other measurements were performed under fed conditions. Wild-type and female mice were euthanized on day 19 of gestation. Blood was centrifuged at 1000 g for 10 min at 4 °C to collect serum and then stored at -80 °C for subsequent analysis. Liver tissue was harvested and stored at -80 °C for biochemical analysis. Total serum cholesterol (TChl), triglycerides (TG), high-density lipoprotein (HDL), low-density lipoprotein (LDL), and Malondialdehyde (MDA) levels were measured using a total cholesterol assay kit, serum triglyceride quantification kit, HDL and LDL cholesterol assay kit, and Lipid Peroxidation MDA Assay Kit. Liver lysates were prepared as previously described. Liver MDA, Superoxide dismutase (SOD), GPx, Glutathione (GSH), and CAT using Lipid Peroxidation MDA Assay Kit, Total SOD Assay Kit and NBT, Total Glutathione Peroxidase Assay Kit, Glutathione reductase detection kit measurement, and catalase detection kit were purchased from Beyotime Biotechnology (Shanghai, China).

THP-1 derived macrophages were divided randomly into four groups: (1) control, (2) ultrasound alone (ultrasound), (3) P-HY alone (P-HY), and (4) P-HY plus ultrasound (SDT). For the P-HY and SDT groups, the cells were incubated with 0.4 µg/mL P-HY for a 4 h drug-loading time in RPMI 1640 medium supplemented with 10% FBS.
For the control and ultrasound groups, an equivalent volume of medium was used to replace P-HY. The cells in the ultrasound and SDT groups were exposed to an ultrasound at a frequency of 1.0 MHz and an intensity of 0.5 W/cm 2 for 15 min. After the treatment, the cells were cultured in fresh medium for a further 6 h and then prepared for different analyses.
For inhibitory studies, 10 mM sodium azide (NaN 3 , Sigma, Aldrich, USA), mannitol (Beyotime Biotechnology, Inc., Beijing, China), 100 µg/mL superoxide dismutase (SOD, Beyotime Biotechnology, Inc., Beijing, China), 100 µg/mL catalase (CAT, Beyotime Biotechnology, Inc., Beijing, China), 1 mM N-acetyl cysteine (NAC, Sigma, Aldrich, USA), 20 µM z-VAD-FMK (z-VAD, BioVision Inc., USA), 0.5 µM cyclosporin A (CsA, Sigma, Aldrich, USA), 100 µM bongkrekic acid (BA, Sigma, Aldrich, USA) and 1 µM 4, 4'-diisothiocyanatostilbene-2,2'-disulfonic acid disodium (DIDS, Sigma, Aldrich, USA) were incubated together with P-HY for 4 h.

Dulbecco's modified Eagle's medium (DMEM) medium and fetal bovine serum (FBS) was purchased from GIBCO-BRL (USA). N-acetyl-L-cysteine (NAC), diphenylene iodonium (DPI), NaHS, and LPS (E. coli serotype 055:B5) were obtained from Sigma (St. Louis, MO). Carboxy-PTIO potassium salt (c-PTIO), catalase (CAT), and superoxide dismutase (SOD) were purchased from Beyotime (Beyotime Biotechnology, Jiangsu, China). LY294002 and SB203580 were obtained from Calbiochem (Dermstadt, Germany). Antibodies against β-Actin, CSE, iNOS, COX-2, Nox4 were obtained from Santa Cruz Biotechnology (Santa Cruz, CA). Total-and phosphor (p) -p38 (Thr 180 and Tyr 182 ), total-and p-extracellular signal-regulated kinase 1/2 (ERK) (Thr 202 / Tyr 204 ), total-and p-Akt (Ser 473 ), and p-glycogen synthase kinase-3β (GSK3β) (Ser 9 ) were purchased from Cell Signaling Technology (Danvers, MA).