Journal: Antioxidants

Article Title: Placental Adaptive Changes to Protect Function and Decrease Oxidative Damage in Metabolically Healthy Maternal Obesity

doi: 10.3390/antiox9090794

Figure Lengend Snippet: NO-induced nitrosative stress markers in placental samples from normal weight (lean, n = 9) and obese (OB, n = 8) pregnancies. ( a ) iNOS expression levels. This enzyme was assessed in placental protein extracts by Western blot analysis, as described in the “Material and Methods” Section. ( b ) Detection and quantification of ERK nitrosylated (SNO-ERK1,2) levels. The placental extracts were processed and subjected to the Biotin-Switch method. The biotin-labeled SNO proteins were further subjected to neutravidin capture, separated by using 10% by SDS-PAGE, and immunoblotted with an anti-ERK antibody. Total ERK were represented as total protein (input). ( c ) Detection of nitrated proteins by immunoblot analysis using antinitrotyrosine antibodies. In each lane, all nitrated proteins were counted in order to quantify the total nitration protein level for each placental sample. Molecular weight markers (M) and sizes (kDa) are shown, and also possible targets of nitrated proteins (arrows) are indicated. In all cases: top represents the mean densitometry expressed in arbitrary units and bottom shows a representative picture of all Western blots. The results were expressed as means ± SD. * p < 0.05 relative to the lean group or input. iNOS: inducible nitric oxide synthase.

Article Snippet: Primary (anti-ERK1/2, anti-SOD 1, anti-catalase, and β-actin) and secondary antibodies came from Cell Signaling Technology (Danvers, MA, USA). iNOS antibody was from R&D Systems (Minneapolis, MN, USA) and nitrotyrosine antibody was from Merck Millipore (Darmstadt, Germany).

Techniques: Expressing, Western Blot, Labeling, SDS Page, Nitration, Molecular Weight