Role of nitric oxide synthase types II and III in early protection against endotoxin-induced lung injury: RESULT (part 6)

Identification of NOS-II transcripts and kinetics of their appearance: Freshly homogenized lung tissue from controls and from LPS-treated rats (decapitated 15, 30 and 120 mins after LPS infusion) were subjected to total RNA isolation by the Trizol method. RNA (1 |ig) was reverse transcribed and amplified by PCR with rat-specific oligonucleotide primers for NOS-II (Figure 5). Beta-actin-specific primers were used as a positive control. Negative control was obtained from the samples in the absence of any primers. A 384 base pair single band was amplified from samples with NOS-II primers 30 and 120 mins after LPS infusion. No RT-PCR NOS-II-specific product was amplified from control tissues or from tissues treated for 15 mins with LPS. The expected 308 base pair band with the beta-actin-specific primers was observed in every tested sample. As early as 30 mins after LPS infusion, the 384 base pair single band product specific for NOS-II was detected by RT-PCR technique. This finding prompted a comparison of the expression of NOS-II mRNA in various tissues of the rat (lung, liver, spleen, kidney and heart) at different periods after LPS infusion by dot blot analysis using PCR-generated specific cDNA probe for NOS-II and 18S RNA probe as a control. buy ampicillin


This entry was posted in Lung injury and tagged 1-Amino-2-hydroxy-guanidine, Endotoxic shock, NG-nitro-L-arginine, Nitric oxide, Nitric oxide synthase type II, Nitric oxide synthase type III, S-nitroso-N-acetyl-penicillamine.