A disease-associated gene desert directs macrophage inflammation through ETS2.

Increasing rates of autoimmune and inflammatory disease present a burgeoning threat to human health1. This is compounded by the limited efficacy of available treatments1 and high failure rates during drug development2, highlighting an urgent need to better understand disease mechanisms. Here we show how functional genomics could address this challenge. By investigating an intergenic haplotype on chr21q22-which has been independently linked to inflammatory bowel disease, ankylosing spondylitis, primary sclerosing cholangitis and Takayasu's arteritis3-6-we identify that the causal gene, ETS2, is a central regulator of human inflammatory macrophages and delineate the shared disease mechanism that amplifies ETS2 expression. Genes regulated by ETS2 were prominently expressed in diseased tissues and more enriched for inflammatory bowel disease GWAS hits than most previously described pathways. Overexpressing ETS2 in resting macrophages reproduced the inflammatory state observed in chr21q22-associated diseases, with upregulation of multiple drug targets, including TNF and IL-23. Using a database of cellular signatures7, we identified drugs that might modulate this pathway and validated the potent anti-inflammatory activity of one class of small molecules in vitro and ex vivo. Together, this illustrates the power of functional genomics, applied directly in primary human cells, to identify immune-mediated disease mechanisms and potential therapeutic opportunities.

High-resolution air-clad imaging fibers.

We present a coherent fiber bundle comprising over 11,000 doped silica cores separated by an air-filled cladding. The fiber is characterized, and its imaging quality is shown to be a substantial improvement over the commercial state of the art, with comparable resolution over an unparalleled spectral range.

Investigation of a possible extended risk haplotype in the IL23R region associated with ankylosing spondylitis.

The IL23R region on chromosome 1 exhibits complex associations with ankylosing spondylitis (AS). We used publicly available epigenomic information and historical genetic association data to identify a putative regulatory element (PRE) in the intergenic region between IL23R and IL12RB2, which includes two single-nucleotide polymorphisms (SNPs) independently associated with AS-rs924080 (P=2 × 10-3) and rs11578380 (P=2 × 10-4). In luciferase reporter assays, this PRE showed silencer activity (P<0.001). Haplotype and conditional analysis of 4230 historical AS cases and 9700 controls revealed a possible AS-associated extended haplotype, including the PRE and risk variants at three SNPs (rs11209026, rs11209032 and rs924080), but excluding the rs11578380 risk variant. However, the rs924080 association was absent after conditioning on the primary association with rs11209032, which, in contrast, was robust to conditioning on all other AS-associated SNPs in this region (P<2 × 10-8). The role of this putative silencer on some IL23R extended haplotypes therefore remains unclear.

miR-10b-5p is a novel Th17 regulator present in Th17 cells from ankylosing spondylitis.

OBJECTIVE: To determine the microRNA (miR) signature in ankylosing spondylitis (AS) T helper (Th)17 cells. METHODS: Interleukin (IL)-17A-producing CD4+ T cells from patients with AS and healthy controls were FACS-sorted for miR sequencing and qPCR validation. miR-10b function was determined by miR mimic expression followed by cytokine measurement, transcriptome analysis, qPCR and luciferase assays. RESULTS: AS Th17 cells exhibited a miR signature characterised by upregulation of miR-155-5p, miR-210-3p and miR-10b. miR-10b has not been described previously in Th17 cells and was selected for further characterisation. miR-10b is transiently induced in in vitro differentiated Th17 cells. Transcriptome, qPCR and luciferase assays suggest that MAP3K7 is targeted by miR-10b. Both miR-10b overexpression and MAP3K7 silencing inhibited production of IL-17A by both total CD4 and differentiating Th17 cells. CONCLUSIONS: AS Th17 cells have a specific miR signature and upregulate miR-10b in vitro. Our data suggest that miR-10b is upregulated by proinflammatory cytokines and may act as a feedback loop to suppress IL-17A by targeting MAP3K7. miR-10b is a potential therapeutic candidate to suppress pathogenic Th17 cell function in patients with AS.

Chronic mucocutaneous candidiasis: characterization of a family with STAT-1 gain-of-function and development of an ex-vivo assay for Th17 deficiency of diagnostic utility.

Chronic mucocutaneous candidiasis (CMC) is characterized by recurrent and persistent superficial infections, with Candida albicans affecting the mucous membranes, skin and nails. It can be acquired or caused by primary immune deficiencies, particularly those that impair interleukin (IL)-17 and IL-22 immunity. We describe a single kindred with CMC and the identification of a STAT1 GOF mutation by whole exome sequencing (WES). We show how detailed clinical and immunological phenotyping of this family in the context of WES has enabled revision of disease status and clinical management. Together with analysis of other CMC cases within our cohort of patients, we used knowledge arising from the characterization of this family to develop a rapid ex-vivo screening assay for the detection of T helper type 17 (Th17) deficiency better suited to the routine diagnostic setting than established in-vitro techniques, such as intracellular cytokine staining and enzyme-linked immunosorbent assay (ELISA) using cell culture supernatants. We demonstrate that cell surface staining of unstimulated whole blood for CCR6⁺ CXCR3⁻ CCR4⁺ CD161⁺ T helper cells generates results that correlate with intracellular cytokine staining for IL-17A, and is able to discriminate between patients with molecularly defined CMC and healthy controls with 100% sensitivity and specificity within the cohort tested. Furthermore, removal of CCR4 and CD161 from the antibody staining panel did not affect assay performance, suggesting that the enumeration of CCR6⁺ CXCR3⁻ CD4⁺ T cells is sufficient for screening for Th17 deficiency in patients with CMC and could be used to guide further investigation aimed at identifying the underlying molecular cause.

A functional AT/G polymorphism in the 5'-untranslated region of SETDB2 in the IgE locus on human chromosome 13q14.

The immunoglobulin E (IgE)-associated locus on human chromosome 13q14 influencing asthma-related traits contains the genes PHF11 and SETDB2. SETDB2 is located in the same linkage disequilibrium region as PHF11 and polymorphisms within SETDB2 have been shown to associate with total serum IgE levels. In this report, we sequenced the 15 exons of SETDB2 and identified a single previously ungenotyped mutation (AT/G, rs386770867) in the 5'-untranslated region of the gene. The polymorphism was found to be significantly associated with serum IgE levels in our asthma cohort (P=0.0012). Electrophoretic mobility shift assays revealed that the transcription factor Ying Yang 1 binds to the AT allele, whereas SRY (Sex determining Region Y) binds to the G allele. Allele-specific transcription analysis (allelotyping) was performed in 35 individuals heterozygous for rs386770867 from a panel of 200 British families ascertained through probands with severe stage 3 asthma. The AT allele was found to be significantly overexpressed in these individuals (P=1.26×10(-21)). A dual-luciferase assay with the pGL3 luciferase reporter gene showed that the AT allele significantly affects transcriptional activities. Our results indicate that the IgE-associated AT/G polymorphism (rs386770867) regulates transcription of SETDB2.

Highly birefringent 98-core fiber.

We report on the fabrication and characterization of a polarization-maintaining multicore fiber. The fiber has 98 aligned cores, each with a birefringence of ≈2.3×10-4. The beat length, polarization extinction ratio, and polarization orientation are characterized.

Femtosecond pulses at 20 GHz repetition rate through spectral masking of a phase modulated signal and nonlinear pulse compression.

We present a laser system capable of producing 190 femtosecond pulses at a repetition rate of 20 GHz. The spectral masking of a phase modulated diode laser is used to produce a train of picosecond pulses which are compressed using a fibre-grating compressor followed by subsequent adiabatic soliton compression to the femtosecond regime using a tapered photonic crystal fiber.

1064 nm laser-induced defects in pure SiO2 fibers.

We investigate evidence of the formation of nonbridging oxygen hole centers in pure silica photonic crystal fibers from 5 ps 1064 nm pulses. The formation of the defects is attributed to the breaking of stressed silicon-oxygen bonds in the glass matrix through a many-photon process. We compare the photodarkening induced by the 1064 nm pump with photodarkening induced by short wavelength light in a 1064 nm pumped supercontinuum extending to 400 nm. It is shown that the higher peak power at the pump wavelength makes it a more significant cause of photodarkening when compared to the shorter wavelength light generated in the fiber.

A polymorphism that affects OCT-1 binding to the TNF promoter region is associated with severe malaria.

Genetic variation in cytokine promoter regions is postulated to influence susceptibility to infection, but the molecular mechanisms by which such polymorphisms might affect gene regulation are unknown. Through systematic DNA footprinting of the TNF (encoding tumour necrosis factor, TNF) promoter region, we have identified a single nucleotide polymorphism (SNP) that causes the helix-turn-helix transcription factor OCT-1 to bind to a novel region of complex protein-DNA interactions and alters gene expression in human monocytes. The OCT-1-binding genotype, found in approximately 5% of Africans, is associated with fourfold increased susceptibility to cerebral malaria in large case-control studies of West African and East African populations, after correction for other known TNF polymorphisms and linked HLA alleles.

Resolving the variable genome and epigenome in human disease.

The individual human genome and epigenome are being defined at unprecedented resolution by current advances in sequencing technologies with important implications for human disease. This review uses examples relevant to clinical practice to illustrate the functional consequences of genetic and epigenetic variation. The insights gained from genome-wide association studies are described together with current efforts to understand the role of rare variants in common disease, set in the context of recent successes in Mendelian traits through the application of whole exome sequencing. The application of functional genomics to interrogate the genome and epigenome, build up an integrated picture of the regulatory genomic landscape and inform disease association studies is discussed, together with the role of expression quantitative trait mapping and analysis of allele-specific gene expression.

Delivery of high energy Er:YAG pulsed laser light at 2.94 µm through a silica hollow core photonic crystal fibre.

In this paper the delivery of high power Er:YAG laser pulses through a silica hollow core photonic crystal fibre is demonstrated. The Er:YAG wavelength of 2.94 µm is well beyond the normal transmittance of bulk silica but the unique hollow core guidance allows silica to guide in this regime. We have demonstrated for the first time the ability to deliver high energy pulses through an all-silica fibre at 2.94 µm. These silica fibres are mechanically and chemically robust, biocompatible and have low sensitivity to bending. A maximum pulse energy of 14 mJ at 2.94 µm was delivered through the fibre. This, to our knowledge, is the first time a silica hollow core photonic crystal fibre has been shown to transmit 2.94 µm laser light at a fluence exceeding the thresholds required for modification (e.g. cutting and drilling) of hard biological tissue. Consequently, laser delivery systems based on these fibres have the potential for the realization of novel, minimally-invasive surgical procedures.

Modulational instability in a silicon-on-insulator directional coupler: role of the coupling-induced group velocity dispersion.

We report frequency conversion experiments in silicon-on-insulator (SOI) directional couplers. We demonstrate that the evanescent coupling between two subwavelength SOI waveguides is strongly dispersive and significantly modifies modulational instability (MI) spectra through the coupling induced group velocity dispersion (GVD). As the separation between two 380-nm-wide silicon photonic wires decreases, the increasing dispersion of the coupling makes the GVD in the symmetric supermode more normal and suppresses the bandwidth of the MI gain observed for larger separations.

Transcriptional repression and DNA looping associated with a novel regulatory element in the final exon of the lymphotoxin-ß gene.

Transcriptional regulation has a critical role in the coordinate and context-specific expression of a cluster of genes encoding members of the tumour necrosis factor (TNF) superfamily found at chromosome 6p21, comprising TNF, LTA (encoding lymphotoxin-α) and LTB (encoding lymphotoxin-ß). This is important, as dysregulated expression of these genes is implicated in susceptibility to many autoimmune, inflammatory and infectious diseases. We describe here a novel regulatory element in the fourth exon of LTB, which is highly conserved, localises to the only CpG island in the locus, and is associated with a DNase I hypersensitive site and specific histone modifications. We find evidence of binding by Yin Yang 1 (YY1), cyclic AMP response element (CRE)-binding protein (CREB) and CCCTC-binding factor (CTCF) to this region in Jurkat T cells, which is associated with transcriptional repression on reporter gene analysis. Chromatin conformation capture experiments show evidence of DNA looping, involving interaction of this element with the LTB promoter, LTA promoter and TNF 3' untranslated region (UTR). Small interfering RNA (siRNA) experiments demonstrate a functional role for YY1 and CREB in LTB expression. Our findings provide evidence of additional complexity in the transcriptional regulation of LTB with implications for coordinate expression of genes in this important genomic locus.

Coherent supercontinuum generation in photonic crystal fiber with all-normal group velocity dispersion.

We demonstrate supercontinuum generation in a photonic crystal fiber with all-normal group velocity dispersion. Pumping a short section of this fiber with compressed pulses from a compact amplified fiber laser generates a 200 nm bandwidth continuum with typical self-phase-modulation characteristics. We demonstrate that the supercontinuum is compressible to a duration of 26 fs. It therefore has a high degree of coherence between all the frequency components, and is a single pulse in the time domain. A smooth, flat spectrum spanning 800 nm is achieved using a longer piece of fiber.

Compact, stable and efficient all-fibre gas cells using hollow-core photonic crystal fibres.

Gas-phase materials are used in a variety of laser-based applications--for example, in high-precision frequency measurement, quantum optics and nonlinear optics. Their full potential has however not been realized because of the lack of a suitable technology for creating gas cells that can guide light over long lengths in a single transverse mode while still offering a high level of integration in a practical and compact set-up or device. As a result, solid-phase materials are still often favoured, even when their performance compares unfavourably with gas-phase systems. Here we report the development of all-fibre gas cells that meet these challenges. Our structures are based on gas-filled hollow-core photonic crystal fibres, in which we have recently demonstrated substantially enhanced stimulated Raman scattering, and which exhibit high performance, excellent long-term pressure stability and ease of use. To illustrate the practical potential of these structures, we report two different devices: a hydrogen-filled cell for efficient generation of rotational Raman scattering using only quasi-continuous-wave laser pulses; and acetylene-filled cells, which we use for absolute frequency-locking of diode lasers with very high signal-to-noise ratios. The stable performance of these compact gas-phase devices could permit, for example, gas-phase laser devices incorporated in a 'credit card' or even in a laser pointer.

Regulation of major histocompatibility complex class II gene expression, genetic variation and disease.

Major histocompatibility complex (MHC) class II molecules are central to adaptive immune responses and maintenance of self-tolerance. Since the early 1970s, the MHC class II region at chromosome 6p21 has been shown to be associated with a remarkable number of autoimmune, inflammatory and infectious diseases. Given that a full explanation for most MHC class II disease associations has not been reached through analysis of structural variation alone, in this review we examine the role of genetic variation in modulating gene expression. We describe the intricate architecture of the MHC class II regulatory system, indicating how its unique characteristics may relate to observed associations with disease. There is evidence that haplotype-specific variation involving proximal promoter sequences can alter the level of gene expression, potentially modifying the emergence and expression of key phenotypic traits. Although much emphasis has been placed on cis-regulatory elements, we also examine the role of more distant enhancer elements together with the evidence of dynamic inter- and intra-chromosomal interactions and epigenetic processes. The role of genetic variation in such mechanisms may hold profound implications for susceptibility to common disease.

Time and frequency domain measurements of solitons in subwavelength silicon waveguides using a cross-correlation technique.

We report time domain measurements of the group-velocity-dispersion-induced and nonlinearity-induced chirping of femtosecond pulses in subwavelength silicon-on-insulator waveguides. We observe that at a critical input power level, these two effects compensate each other leading to soliton formation. Formation of the fundamental optical soliton is observed at a peak power of a few Watts inside the waveguide. Interferometric cross-correlation traces reveal compression of the soliton pulses, while spectral measurements show pronounced dispersive waves emitted by solitons into the wavelength range of normal group velocity dispersion.

Ultrashort pulse compression and delivery in a hollow-core photonic crystal fiber at 540 nm wavelength.

We have fabricated a bandgap-guiding hollow-core photonic crystal fiber (PCF) capable of transmitting and compressing ultrashort pulses in the green spectral region around 532 nm. When propagating subpicosecond pulses through 1 m of this fiber, we have observed soliton-effect temporal compression by up to a factor of 3 to around 100 fs. This reduces the wavelength at which soliton effects have been observed in hollow-core PCF by over 200 nm. We have used the pulses delivered at the output of the fiber to machine micrometer-scale features in copper.