Figure 1: iNOS is increased and miR-323-5p is decreased in X-ALD patient-derived lymphocytes in vitro. (A) Nitrite (NO) levels were increased in the supernatant of X-ALD patient-derived lymphocytes maintained in serum-free medium for 24-72 h. (B) Quantitative RT-PCR analysis showed that iNOS mRNA expression was increased in X-ALD patient-derived lymphocytes. (C) Quantitative RT-PCR of the same samples for miR-323-5p showed a decreased expression. 20 ng cDNA per sample was amplified by quantitative RT-PCR for mRNA or miRNA analysis. Gene expression was normalized to L27 expression for mRNA and RNU6 for miRNA expression. (D) Viability of lymphocytes maintained in the serum-free media was checked by trypan blue dye exclusion. Data are represented as mean ± SD of three different experiments. ***p<0.001. iNOS: inducible nitric oxide synthase; CTL: healthy control lymphocytes; ALD: X-ALD patient-derived lymphocytes.