Fine-grained all-fiber nonlocal dispersion compensation in the telecommunications O-band.

Nonlocal dispersion compensation between broadband nondegenerate photon pairs propagated over fiber corresponding to the ITU-T G.652D telecommunications standard was studied extensively via fine-grained measurements of the temporal correlation between them. We demonstrated near-ideal levels of nonlocal dispersion compensation by adjusting the propagation distance of the photon pairs to preserve photon timing correlations close to the effective instrument resolution of our detection apparatus (41.0±0.1ps). Experimental data indicates that this degree of compensation can be achieved with relatively large fiber increments (1km), compatible with real-world deployment. Ultimately, photon timing correlations were preserved down to 51ps±21ps over two multi-segmented 10km spans of deployed metropolitan fiber.

Functional and Multi-Omics Effects of an Optimized CRISPR-Mediated FURIN Depletion in U937 Monocytes.

The pro-protein convertase FURIN (PCSK3) is implicated in a wide range of normal and pathological biological processes such as infectious diseases, cancer and cardiovascular diseases. Previously, we performed a systemic inhibition of FURIN in a mouse model of atherosclerosis and demonstrated significant plaque reduction and alterations in macrophage function. To understand the cellular mechanisms affected by FURIN inhibition in myeloid cells, we optimized a CRISPR-mediated gene deletion protocol for successfully deriving hemizygous (HZ) and nullizygous (NZ) FURIN knockout clones in U937 monocytic cells using lipotransfection-based procedures and a dual guide RNA delivery strategy. We observed differences in monocyte and macrophage functions involving phagocytosis, lipid accumulation, cell migration, inflammatory gene expression, cytokine release patterns, secreted proteomics (cytokines) and whole-genome transcriptomics between wild-type, HZ and NZ FURIN clones. These studies provide a mechanistic basis on the possible roles of myeloid cell FURIN in cardiovascular disorders.

An optimized method for gene knockdown in differentiating human and mouse adipocyte cultures.

Adipocyte cultures are a mainstay of metabolic disease research, yet loss-of-function studies in differentiating adipocytes is complicated by the refractoriness of lipid-containing adipocytes to standard siRNA transfections. Alternative methods, such as electroporation or adenovirus/lentivirus-based delivery systems are complex, expensive and often accompanied with unacceptable levels of cell death. To address this problem, we have tested two commercially available siRNA delivery systems in this study using a multi-parameter optimization approach. Our results identified a uniform siRNA transfection protocol that can be applied to human and mouse adipocyte cultures throughout the time course of differentiation, beginning with pre-differentiated cells and continuing up to lipid-accumulated differentiated adipocytes. Our findings allow for efficient transfection of human and mouse adipocyte cultures using standard and readily available methodologies, and should help significantly expand the scope of gene manipulation studies in these cell types.

Interactome Analysis of Visceral Adipose Tissue Elucidates Gene Regulatory Networks and Novel Gene Candidates in Obesity.

Objective: Visceral adiposity is associated with increased proinflammatory activity, insulin resistance, diabetes risk and mortality rate. Numerous individual genes have been associated with obesity, but studies investigating gene-regulatory networks in human visceral obesity are lacking. Methods: We analyzed gene-regulatory networks in human visceral adipose tissue (VAT) from 48 obese and 11 non-obese Chinese subjects using gene co-expression and network construction with RNA-sequencing data. We also conducted RNA interference-based tests on selected genes for adipocyte differentiation effects. Results: A scale-free gene co-expression network was constructed from 360 differentially expressed genes between obese and non-obese VAT (absolute log fold-change >1, FDR<0.05) with edge probability >0.8. Gene regulatory network analysis identified candidate transcription factors associated with differentially expressed genes. Fifteen subnetworks (communities) displayed altered connectivity patterns between obese and non-obese networks. Genes in pro-inflammatory pathways showed increased network connectivities in obese VAT whereas the oxidative phosphorylation pathway displayed reduced connections (enrichment FDR<0.05). Functional screening via RNA interference identified SOX30 and OSBPL3 as potential network-derived gene candidates influencing adipocyte differentiation. Conclusions: This interactome-based approach highlights the network architecture, identifies novel candidate genes, and leads to new hypotheses regarding network-assisted gene regulation in obese vs. non-obese VAT.What is already known about this subject?: Visceral adipose tissue (VAT) is associated with increased levels of proinflammatory activity, insulin resistance, diabetes risk and mortality rate.Gene expression studies have identified candidate genes associated with proinflammatory function in VAT.What are the new findings in your manuscript?: Using integrative network-science, we identified co-expression and gene regulatory networks that are differentially regulated in VAT samples from subjects with and without obesityWe used functional testing (adipocyte differentiation) to validate a subset of novel candidate genes with minimal prior reported associations to obesityHow might your results change the direction of research or the focus of clinical practice: Network biology-based investigation provides a new avenue to our understanding of gene function in visceral adiposityFunctional validation screen allows for the identification of novel gene candidates that may be targeted for the treatment of adipose tissue dysfunction in obesity.