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. 2023 Jun 9;17:1180308. doi: 10.3389/fnins.2023.1180308

Figure 6.

Figure 6

Quantitative real-time PCR validated dysregulated levels of circRNAs and their linear isoforms. (A) Schematic representation of an example circRNA biogenesis resulting from back-splicing events of Exon 2 and Exon 4 from a hypothetical precursor mRNA. The divergent primers for circRNA qRT-PCR validation were designed such that only the back-spliced and not linear junction was amplified. (B) Spinal circRps6ka3 and linear Rps6ka3 levels were measured. A main effect of PAE (F1,31 = 5.53, p = 0.025) was observed in circRps6ka3 levels. An unpaired t-test revealed a significant downregulation of circRps6ka3 levels (p < 0.03) between PAE vs. Sac control, Sham injured rats. mRNA levels were not different across treatment groups. (C) CircItch levels were downregulated with PAE, a main effect (F1,29 = 4.58, p = 0.004) was observed. A main effect of surgery (F1,29 = 15.66, p < 0.05) was also observed. Post-doc test revealed a significant difference between PAE to Sac control (p < 0.005), Sham-injured groups. Although the levels were not significantlyFIGURE 6 (Continued) different with 2-way ANOVA, an unpaired t-test revealed significant downregulation of circItch levels (p < 0.05) between nerve-injured PAE vs. nerve injure Sac control rats. mRNA levels were not different across treatment groups. (D) Blood leukocytes from additional adult Sac (N = 6) and PAE (N = 8) female rats were used for validation. A significant decrease of both circVopp1 (p < 0.01) and linear Vopp1 (p < 0.02) levels was observed in PAE. (E) A main effect of PAE (F1,25 = 4.25, *p < 0.05) was observed on circVopp1 levels in the spinal cord, however, multiple comparisons were not significant between groups. mRNA levels were not different across treatment groups, however, a trend of increased Vopp1 mRNA levels in nerve-injured PAE rats was observed compared to the nerve-injured Sac rats. *Denotes p values with significance.