Hence, the theory that cells subjected to hypoxia release potential TLR ligands is not new

and WT mice As shown in Fig 6, the deletion of ASMase significantly increased SM levels in the retina at various ages (Fig 6A). In addition, several very long chain SMs (C20, C22, C22:1, C24:1, C26, C26:1) displayed large age-dependent increases. Interestingly, sphingosine and dihydrosphingosine levels were also elevated in the ASMase KO mice in an age-dependent manner (Fig 6B). However, no significant changes in the ceramide level were observed between ASMase KO and WT retinas (Fig 6C). Retinal S1P levels were below the level of detection. Autophagy dysfunction in ASMase KO eyecups Autophagy is an intracellular lysosomal clearance process associated with retinal degeneration and sphingolipid disorders [33�35]. As shown in Fig 7A, deletion of ASMase resulted in a 58 �12.8% increase in LC3-II in Scutellarein 19667145?ordinalpos=7&itool=EntrezSystem2.PEntrez.Pubmed.Pubmed_ResultsPanel.Pubmed_DefaultReportPanel.Pubmed_RVDocSum” title=View Abstract(s)”>PubMed ID:http://www.ncbi.nlm.nih.gov/pubmed/19667145 the eyecups, suggesting the accumulation of autophagosomes in ASMase KO eyecups. However, LC3-I and beclin-1, PubMed ID:http://www.ncbi.nlm.nih.gov/pubmed/19668191 the upstream regulators of autophagy signaling and crucial to the initiation of the formation of autophagosomes, showed no measureable difference between WT and ASMase KO eyecups. To determine if the autophagy disorder is a consequence caused by the ASMase deletion, we evaluated the levels of LC-3 isoforms in cultured cells, including a control fibroblast cell line (GM02226) and a Niemann-Pick type A dermal fibroblast cell line (NPD GM13205). As shown in Fig 7B, significant accumulation of LC-3II was observed in the NPD fibroblast cells, while no changes in LC3-I were observed. The accumulation of LC3-II could be caused by either the activation of autophagocytosis or by defects in the fusion process of autophagosomes with lysosomes [36]. To discriminate between these two possibilities, the cells were treated with two lysos