Figure 2. Characterization of ankyrin repeat and suppressor of cytokine signaling (SOCS) box containing protein-10-stained structures. A: The area of ankyrin repeat and suppressor of cytokine signaling (SOCS) box containing protein-10 (ASB10)-stained structures was measured using FIJI software from 25 confocal images of human trabecular meshwork (HTM) cells derived from four individuals. One microscope field shows examples of a large structure and small vesicles. Scale bar=20 µm. B: Efficacy of ASB10 knockdown by sh177 lentivirus was determined. Quantitative PCR (qRT–PCR) was used to measure ASB10 mRNA levels in HTM cells infected with 10⁶ pfu of sh177 or shControl (shCtrl) lentivirus. Values were normalized for 18S RNA. Values are presented as a percentage of the control±SEM; n=3; * p=0.0001 with ANOVA. Western immunoblot analysis shows significant knockdown of the ASB10 protein in the HTM cells. C: The sh177 ASB10-silencing lentivirus was applied to human anterior segment perfusion culture at time point 0, and outflow rate was monitored for a further 93 h. There was a significant (*p<0.001) decrease in outflow rate compared to the control-infected eyes. Error bars are SEM. D: The number of large (>5 µm diameter) ASB10-stained structures per nuclei in the control HTM and ASB10-silenced HTM cells was counted. Error bars are the SEM. *, p=0.033 and **, p=0.03 compared to control short, hairpin ribonucleic acid (shRNA)-infected HTM cells with ANOVA. E: Poisson probability distribution of the large ASB10-stained structures in control and ASB10-silenced HTM cells is shown.