93 research outputs found

    Dysregulation of lipid metabolism in the liver of Tspo knockout mice

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    The translocator protein, TSPO, has been implicated in a wide range of cellular processes exerted from its position in the outer mitochondrial membrane from where it influences lipid metabolism and mitochondrial oxidative activity. Understanding how this protein regulates a profusion of processes requires further elucidation and to that end we have examined lipid metabolism and used an RNAseq strategy to compare transcript abundance in wildtype and Tspo knockout (KO) mouse liver. The levels of cholesterol, triglyceride and phospholipid were significantly elevated in the KO mouse liver. The expression of cholesterol homeostasis genes was markedly downregulated. Determination of the differential expression revealed that many genes were either up- or downregulated in the KO animals. However, a striking observation within the results was a decrease of transcripts for protein degradation proteins in KO animals while protease inhibitors were enriched. When the entire abundance data-set was analysed with CEMiTool, and revealed a module of proteins that were under-represented in the KO animals. These could subsequently be formed into a network comprising three interlinked clusters at the centre of which were proteins of cytoplasmic ribosomes with gene ontology terms suggesting impairment to translation. The largest cluster was dominated by proteins of lipid metabolism but also contained disparate systems of iron metabolism and behaviour. The third cluster was dominated by proteins of the electron transport chain and oxidative phosphorylation. These findings suggest that TSPO contributes to lipid metabolism, detoxification of active oxygen species and oxidative phosphorylation, and regulates mitochondrial retrograde signalling

    Deletion of TSPO causes dysregulation of cholesterol metabolism in mouse retina

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    Cholesterol dysregulation has been implicated in age-related macular degeneration (AMD), the most common cause of visual impairment in the elderly. The 18 KDa translocator protein (TSPO) is a mitochondrial outer membrane protein responsible for transporting cholesterol from the mitochondrial outer membrane to the inner membrane. TSPO is highly expressed in retinal pigment epithelial (RPE) cells, and TSPO ligands have shown therapeutic potential for the treatment of AMD. Here, we characterized retinal pathology of Tspo knockout (KO) mice using histological, immunohistochemical, biochemical and molecular biological approaches. We found that Tspo KO mice had normal retinal morphology (by light microscopy) but showed elevated levels of cholesterol, triglycerides and phospholipids with perturbed cholesterol efflux in the RPE cells of Tspo KO mice. Expression of cholesterol-associated genes (Nr1h3, Abca1, Abcg1, Cyp27a1 and Cyp46a1) was significantly downregulated, and production of pro-inflammatory cytokines was markedly increased in Tspo KO retinas. Furthermore, microglial activation was also observed in Tspo KO mouse retinas. These findings provide new insights into the function of TSPO in the retina and may aid in the design of new therapeutic strategies for the treatment of AMD

    Chinese medicine, Qijudihuang pill, mediates cholesterol metabolism and regulates gut microbiota in high-fat diet-fed mice, implications for age related macular degeneration

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    Background: Traditional Chinese Medicines have been used for thousands of years but without any sound empirical basis. One such preparation is the Qijudihuang pill (QP), a mixture of eight herbs, that has been used in China for the treatment of various conditions including age-related macular degeneration (AMD), the most common cause of blindness in the aged population. In order to explain the mechanism behind the effect of QP, we used an AMD model of high-fat diet (HFD) fed mice to investigate cholesterol homeostasis, oxidative stress, inflammation and gut microbiota.Methods: Mice were randomly divided into three groups, one group was fed withcontrol diet (CD), the other two groups were fed with high-fat-diet (HFD). OneHFD group was treated with QP, both CD and the other HFD groups were treatedwith vehicles. Tissue samples were collected after the treatment. Cholesterollevels in retina, retinal pigment epithelium (RPE), liver and serum weredetermined using a commercial kit. The expression of enzymes involved incholesterol metabolism, inflammation and oxidative stress was measured withqRT-PCR. Gut microbiota was analyzed using 16S rRNA sequencing.Results: In the majority of the lipid determinations, analytes were elevated by HFD but thiswas reversed by QP. Cholesterol metabolism including the enzymes of bile acid (BA) formationwas suppressed by HFD but again thiswas reversed by QP. BAs play amajor role in signaling between host andmicrobiome and this is disrupted by HFD resulting in major changes in the composition of colonic bacterial communities. Associated with these changes are predictions of the metabolic pathway complexity and abundance of individual pathways. These concerned substrate breakdowns, energy production and the biosynthesis of proinflammatory factors but were changed back to control characteristics by QP.Conclusion: We propose that the ability of QP to reverse these HFD-inducedeffects is related to mechanisms acting to lower cholesterol level, oxidative stress and inflammation, and to modulate gut microbiota

    Array-based sequencing of filaggrin gene for comprehensive detection of disease-associated variants

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    The filaggrin gene (FLG) is essential for skin differentiation and epidermal barrier formation. FLG loss-of-function (LoF) variants are associated with ichthyosis vulgaris and the major genetic risk factor for developing atopic dermatitis (AD).1, 2, 3 Genetic stratification of patients with AD according to FLG LoF risk is a common practice for both research and clinical studies; however, few studies comprehensively sequence the entire FLG coding region. Most studies that include FLG genotyping have screened for common predominant LoF variants to report allele frequencies after full Sanger sequencing of a smaller batch of test patient samples or previously published data. This strategy potentially results in underreporting of the genetic contribution especially in ethnicities where FLG LoF variants are highly diverse.4 Distinct LoF variants have been reported for most ethnicities studied to date. For example, 2 predominant sequence variants (p.R501X and c.2282del4) make up approximately 80% of the mutation burden in northern Europeans,5 whereas in East Asian ethnicities, a larger FLG LoF mutation spectrum is found with fewer predominating variants.6, 7 However, routinely Sanger sequencing the entire FLG coding region for large cohorts is not always feasible, although desirable as it is essential to correctly stratify patients. To address this, we developed a robust and cost-effective high-throughput PCR-based method for analyzing the entire coding region of FLG using Fluidigm microfluidics technology and next-generation sequencing (NGS). We have applied this method to fully resequence cohorts of Chinese, Malay, and Indian patients with AD from the Singaporean population.ASTAR (Agency for Sci., Tech. and Research, S’pore)Published versio

    Yorba Times: Standing Up, Speaking Out

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    During the Spring 2018 semester, Dr. Noah Asher Golden\u27s Teaching of Writing K-12 students partnered with the Journalism class at Yorba Academy for the Arts. Through collaboration over a four-month period, Chapman\u27s future teachers and Yorba\u27s junior high journalists engaged a deep writing process to write a series of features, editorials, and news articles related to a number of global issues. Thank you to Ms. Andrea Lopez, Ms. Kori Shelton, Mr. Nick Sepulveda, Ms. Tracy Knibb, and the Lloyd E. and Elisabeth H. Klein Family Foundation for supporting this project.https://digitalcommons.chapman.edu/yorba-chapman/1003/thumbnail.jp
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