Myeloid C/EBP beta deficiency reshapes microglial gene expression and is protective in experimental autoimmune encephalomyelitis
JOURNAL OF NEUROINFLAMMATION
Authors: Pulido-Salgado, Marta; Vidal-Taboada, Jose M.; Diaz-Barriga, Gerardo Garcia; Serratosa, Joan; Valente, Tony; Castillo, Paola; Matalonga, Jonathan; Straccia, Marco; Canals, Josep M.; Valledor, Annabel; Sola, Carme; Saura, Josep
Background: CCAAT/enhancer binding protein beta ( C/ EBP beta) is a transcription factor that regulates the expression of important pro-inflammatory genes in microglia. Mice deficient for C/ EBPa show protection against excitotoxic and ischemic CNS damage, but the involvement in this neuroprotective effect of the various C/ EBP beta-expressing cell types is not solved. Since C/ EBP beta-deficient microglia show attenuated neurotoxicity in culture, we hypothesized that specific C/ EBPa deficiency in microglia could be neuroprotective in vivo. In this study, we have tested this hypothesis by generating mice with myeloid C/ EBP beta deficiency. Methods: Mice with myeloid C/ EBP beta deficiency were generated by crossing LysMCre and C/ EBP beta(fl/) (fl) mice. Primary microglial cultures from C/ EBP beta fl/ fl and LysMCre- C/ EBP beta fl mice were treated with lipopolysaccharide interferon. ( IFN gamma.) for 6 h, and gene expression was analyzed by RNA sequencing. Gene expression and C/ EBP beta deletion were analyzed in vivo in microglia isolated from the brains of C/ EBP beta(f1/f1) fl and LysMCre- C/ EBP beta(f1/f1) fl mice treated systemically with lipolysaccharide or vehicle. Mice of LysMCre- C/ EBP beta fl or control genotypes were subjected to experimental autoimmune encephalitis and analyzed for clinical signs for 52days.One- or two-way ANOVA or Kruskal- Wallis with their appropriate post hoc tests were used. Results: LysMCre- C/ EBP beta(f1/f1) fl mice showed an efficiency of C/ EBP beta deletion in microglia of 100 and 90% in vitro and in vivo, respectively. These mice were devoid of female infertility, perinatal mortality and reduced lifespan that are associated to full C/ EBP beta deficiency. Transcriptomic analysis of C/ EBP beta deficient primary microglia revealed C/ EBP beta dependent expression of 1068 genes, significantly enriched in inflammatory and innate immune responses GO terms. In vivo, microglial expression of the pro-inflammatory genes Cybb, Ptges, Il23a, Tnf and Csf3 induced by systemic lipopolysaccharide injection was also blunted by C/ EBP beta deletion. CNS expression of C/ EBP beta was upregulated in experimental autoimmune encephalitis and in multiple sclerosis samples. Finally, LysMCre- C/ EBP beta(F1/F1) mice showed robust attenuation of clinical signs in experimental autoimmune encephalitis. Conclusion: This study provides new data that support a central role for C/ EBP beta in the biology of activated microglia, and it offers proof of concept for the therapeutic potential of microglial C/ EBP beta inhibition in multiple sclerosis.
Novel atherogenic pathways from the differential transcriptome analysis of diabetic epicardial adipose tissue
NUTRITION METABOLISM AND CARDIOVASCULAR DISEASES
Authors: Camarena, V.; Sant, D.; Mohseni, M.; Salerno, T.; Zaleski, M. L.; Wang, G.; Iacobellis, G.
Background and aim: To evaluate the epicardial adipose tissue (EAT) transcriptome in comparison to subcutaneous fat (SAT) in coronary artery disease (CAD) and type 2 diabetes (T2DM). Methods and results: SAT and EAT samples were obtained from subjects with T2DM and CAD (n = 5) and those without CAD with or without T2DM (= 3) undergoing elective cardiac surgery. RNA-sequencing analysis was performed in both EAT and SAT. Gene enrichment analysis was conducted to identify pathways affected by the differentially expressed genes. Changes of top genes were verified by quantitative RT-PCR (qRT-PCR), western blot, and immunofluorescence. A total of 592 genes were differentially expressed in diabetic EAT, whereas there was no obvious changes in SAT transcriptome between diabetics and non-diabetics. Diabetic EAT was mainly enriched in inflammatory genes, such as Colony Stimulating Factor 3 (CSF3), Interleukin-1b (IL-1b), IL-6. KEGG pathway analysis confirmed that upregulated genes were involved in inflammatory pathways, such as Tumor Necrosis Factor (TNF), Nuclear Factor-kappa B (NF-kappa B) and advanced glycation end-products-receptor advanced glycation end products (AGE-RAGE). The overexpression of inflammatory genes in diabetic EAT was largely correlated with upregulated transcription factors such as NF-kappa B and FOS. Conclusions: Diabetic EAT transcriptome is significantly different when compared to diabetic SAT and highly enriched with genes involved in innate immune response and endothelium, like Pentraxin3 (PTX3) and Endothelial lipase G (LIPG). EAT inflammatory genes expression could be induced by upregulated transcription factors, mainly NF-kappa B and FOSL, primarily activated by the overexpressed AGE-RAGE signaling. This suggests a unique and novel atherogenic pathway in diabetes. (C) 2017 The Italian Society of Diabetology, the Italian Society for the Study of Atherosclerosis, the Italian Society of Human Nutrition, and the Department of Clinical Medicine and Surgery, Federico II University. Published by Elsevier B.V. All rights reserved.