Ultra-broadband high-resolution microdroplet spectrometers for the near infrared.

By stabilizing the evaporation dynamics of a microliter fluorocarbon droplet, we demonstrate a fast-scan optofluidic Fourier transform (FT) spectrometer on the tip of an optical fiber operating in the 1000-2000 nm window with a resolution of 3.5 cm-1 (i.e., <1 nm at 1560 nm). Compared with other FT near-infrared (NIR) small-scale spectrometers reported in the literature, the fluorocarbon droplet spectrometer shows the largest wavelength span and span/resolution ratio, allowing spectral analysis of broadband or narrowband radiation to be easily performed. Our results open the way for the practical application of droplet spectrometers as advanced optofluidic NIR analyzers with small size and low cost that are capable of operating in harsh environments, even in the absence of electrical power sources.

Light pressure in droplet micro-resonators excited by free-space scattering.

A droplet optical resonator is a unique environment to investigate light-matter interaction and optomechanics in liquids. Here, we report on light pressure effects derived from whispering gallery modes excited in a liquid-polymer droplet micro-resonator by free-space laser scattering. From the nonlinear resonance spectrum observed in the visible, we provide evidence of photon pressure exerted at the liquid-air boundary and quantify it with a coherent physical model. Our findings pave the way to studies on micro-rheology and nonlinear optofluidics, where droplets serve as miniature liquid laboratories.

A kinetics-based approach to amyloid PET semi-quantification.

PURPOSE: To develop and validate a semi-quantification method (time-delayed ratio, TDr) applied to amyloid PET scans, based on tracer kinetics information. METHODS: The TDr method requires two static scans per subject: one early (~ 0-10 min after the injection) and one late (typically 50-70 min or 90-100 min after the injection, depending on the tracer). High perfusion regions are delineated on the early scan and applied onto the late scan. A SUVr-like ratio is calculated between the average intensities in the high perfusion regions and the late scan hotspot. TDr was applied to a naturalistic multicenter dataset of 143 subjects acquired with [18F]florbetapir. TDr values are compared to visual evaluation, cortical-cerebellar SUVr, and to the geometrical semi-quantification method ELBA. All three methods are gauged versus the heterogeneity of the dataset. RESULTS: TDr shows excellent agreement with respect to the binary visual assessment (AUC = 0.99) and significantly correlates with both validated semi-quantification methods, reaching a Pearson correlation coefficient of 0.86 with respect to ELBA. CONCLUSIONS: TDr is an alternative approach to previously validated ones (SUVr and ELBA). It requires minimal image processing; it is independent on predefined regions of interest and does not require MR registration. Besides, it takes advantage on the availability of early scans which are becoming common practice while imposing a negligible added patient discomfort.

Approaching the transit time limit for high-precision spectroscopy on metastable CO around 6 µm.

As molecular spectroscopy makes its comeback to the limelight of fundamental sciences, scientists need ever better coherent light sources and diagnostic methods. Of particular importance for molecular spectroscopy is the mid infrared spectral region, where strong and narrow ro-vibrational excitations have their fundamental transition frequencies. Unfortunately, much technology in some portions of this spectral region is still rather pioneering. Here we present a high-resolution spectroscopy experiment, based on a molecular beam setup, which pushes the measured linewidth close to the transit time limit, on the order of 100 kHz. Moreover, we discuss the issue of frequency-noise characterization and the linewidth measurement of the ultrastable infrared laser used in the experiment.

Unique chemotactic response profile and specific expression of chemokine receptors CCR4 and CCR8 by CD4(+)CD25(+) regulatory T cells.

Chemokines dictate regional trafficking of functionally distinct T cell subsets. In rodents and humans, a unique subset of CD4(+)CD25(+) cytotoxic T lymphocyte antigen (CTLA)-4(+) regulatory T cells (Treg) has been proposed to control peripheral tolerance. However, the molecular basis of immune suppression and the trafficking properties of Treg cells are still unknown. Here, we determined the chemotactic response profile and chemokine receptor expression of human blood-borne CD4(+)CD25(+) Treg cells. These Treg cells were found to vigorously respond to several inflammatory and lymphoid chemokines. Treg cells specifically express the chemokine receptors CCR4 and CCR8 and represent a major subset of circulating CD4(+) T cells responding to the chemokines macrophage-derived chemokine (MDC)/CCL22, thymus and activation-regulated chemokine (TARC)/CCL17, I-309/CCL1, and to the virokine vMIP-I (ligands of CCR4 and CCR8). Blood-borne CD4(+) T cells that migrate in response to CCL1 and CCL22 exhibit a reduced alloproliferative response, dependent on the increased frequency of Treg cells in the migrated population. Importantly, mature dendritic cells preferentially attract Treg cells among circulating CD4(+) T cells, by secretion of CCR4 ligands CCL17 and CCL22. Overall, these results suggest that CCR4 and/or CCR8 may guide Treg cells to sites of antigen presentation in secondary lymphoid tissues and inflamed areas to attenuate T cell activation.

Differential expression of chemokine receptors and chemotactic responsiveness of type 1 T helper cells (Th1s) and Th2s.

T helper cells type 1 (Th1s) that produce interferon-gamma predominantly mediate cellular immune responses and are involved in the development of chronic inflammatory conditions, whereas Th2s which produce large amounts of IL-4 and IL-5 upregulate IgE production and are prominent in the pathogenesis of allergic diseases. The precise factors determining whether Th1- or Th2-mediated immune responses preferentially occur at a peripheral site of antigen exposure are largely unknown. Chemokines, a superfamily of polypeptide mediators, are a key component of the leukocyte recruitment process. Here we report that among four CXC (CXCR1-4) and five CC (CCR1-5) chemokine receptors analyzed, CXCR3 and CCR5 are preferentially expressed in human Th1s. In contrast, Th2s preferentially express CCR4 and, to a lesser extent, CCR3. In agreement with the differential chemokine receptor expression, Th1s and Th2s selectively migrate in response to the corresponding chemokines. The differential expression of chemokine receptors may dictate, to a large extent, the migration and tissue homing of Th1s and Th2s. It may also determine different susceptibility of Th1s and Th2s to human immunodeficiency virus strains using different fusion coreceptors.

Recruitment and activation of PTP1C in negative regulation of antigen receptor signaling by Fc gamma RIIB1.

Coligation of the Fc receptor on B cells, Fc gamma RIIB1, with the B cell antigen receptor (BCR) leads to abortive BCR signaling. Here it was shown that the Fc gamma RIIB1 recruits the phosphotyrosine phosphatase PTP1C after BCR coligation. This association is mediated by the binding of a 13-amino acid tyrosine-phosphorylated sequence to the carboxyl-terminal Src homology 2 domain of PTP1C and activates PTP1C. Inhibitory signaling and PTP1C recruitment are dependent on the presence of the tyrosine within the 13-amino acid sequence. Inhibitory signaling mediated by Fc gamma RIIB1 is deficient in motheaten mice which do not express functional PTP1C. Thus, PTP1C is an effector of BCR-Fc gamma RIIB1 negative signal cooperativity.

Prognostic Value of 18F-Fluorocholine PET Parameters in Metastatic Castrate-Resistant Prostate Cancer Patients Treated with Docetaxel.

Background and Aim: The availability of new treatments for metastatic castrate-resistant prostate cancer (mCRPC) patients increases the need for reliable biomarkers to help clinicians to choose the better sequence strategy. The aim of the present retrospective and observational work is to investigate the prognostic value of 18F-fluorocholine (18F-FCH) positron emission tomography (PET) parameters in mCRPC. Materials and Methods: Between March 2013 and August 2016, 29 patients with mCRPC were included. They all received three-weekly docetaxel after androgen deprivation therapy, and they underwent 18F-FCH PET/computed tomography (CT) before and after the therapy. Semi-quantitative indices such as maximum standardized uptake value (SUVmax), mean standardized uptake value (SUVmean) with partial volume effect (PVC-SUV) correction, metabolically active tumour volume (MATV), and total lesion activity (TLA) with partial volume effect (PVC-TLA) correction were measured both in pre-treatment and post-treatment 18F-FCH PET/CT scans for each lesion. Whole-body indices were calculated as sum of values measured for each lesion (SSUVmax, SPVC-SUV, SMATV, and STLA). Progression-free survival (PFS) and overall survival (OS) were considered as clinical endpoints. Univariate and multivariate hazard ratios for whole-body 18F-FCH PET indices were performed, and p < 0.05 was considered as significant. Results: Cox regression analysis showed a statistically significant correlation between PFS, SMATV, and STLA. No correlations between OS and 18F-FCH PET parameters were defined probably due to the small sample size. Conclusions: Semi-quantitative indices such as SMATV and STLA at baseline have a prognostic role in patients treated with docetaxel for mCRPC, suggesting a potential role of 18F-FCH PET/CT imaging in clinical decision-making.

Beta2-agonists prevent Th1 development by selective inhibition of interleukin 12.

Interleukin 12 (IL-12) plays a central role in the immune system by skewing the immune response towards T helper 1 (Th1) type responses which are characterized by high interferon-gamma and low IL-4 production. In this report we present evidence that beta2-agonists inhibit IL-12 production by both human monocytes in response to lipopolysaccharide (LPS) and dendritic cells stimulated via CD40. Inhibition of IL-12 production is selective, as other cytokines produced by monocytes are unaffected. IL-12 inhibition is dependent on beta2-adrenoceptor stimulation and correlates with increased levels of intracellular cAMP. In conjunction with their ability to suppress IL-12 production, when beta2-agonists are added at priming of neonatal T lymphocytes, they inhibit the development of Th1-type cells, while promoting T helper 2 (Th2) cell differentiation. Further, the in vivo administration of a therapeutic dose of salbutamol results in the selective inhibition of IL-12 production by whole blood lymphocytes stimulated in vitro with LPS. These findings provide new insight into the immunological consequences of the clinical use of beta2-agonists and may suggest new approaches for the treatment of Th1-mediated diseases.

Fractalkine (CX3CL1) as an amplification circuit of polarized Th1 responses.

Fractalkine (FKN, CX3CL1) is a membrane-bound CX3C chemokine induced by primary proinflammatory signals in vascular endothelial cells (ECs). Here we examined the role of FKN in polarized Th1 or Th2 responses. Proinflammatory signals, including LPS, IL-1, TNF, and CD40 ligand, induced FKN, as did IFN-gamma, which had synergistic activity with TNF. IL-4 and IL-13 did not stimulate the expression of FKN and markedly reduced induction by TNF and IFN-gamma. TNF alone or combined with IFN-gamma also induced release of soluble FKN, which was inhibited by IL-4 and IL-13. In light of this differential regulation of FKN by the master cytokines that control polarized responses, we analyzed the interaction of FKN with natural killer (NK) cells and polarized T-cell populations. NK cells expressed high levels of the FKN receptor CX3CR1 and responded to FKN. CX3CR1 was preferentially expressed in Th1 compared with Th2 cells. Th1 but not Th2 cells responded to FKN. By immunohistochemistry, FKN was expressed on ECs in psoriasis, a Th1-dominated skin disorder, but not in Th2-driven atopic dermatitis. Similarly, ECs in Mycobacterium tuberculosis granulomatous lymphadenitis, but not those in reactive lymph node hyperplasia or in Castelman's disease, showed immunoreactive FKN. These results indicate that regulated expression of FKN in ECs participates in an amplification circuit of polarized type I responses.

Inhibition of IL-12 production by 1,25-dihydroxyvitamin D3. Involvement of NF-kappaB downregulation in transcriptional repression of the p40 gene.

Interleukin 12 (IL-12), produced by myelomonocytic cells, plays a pivotal role in the development of T helper 1 (Th1) cells, which are involved in the pathogenesis of chronic inflammatory autoimmune disorders. 1,25-Dihydroxyvitamin D3 [1,25(OH)2D3] inhibits IL-12 production by activated macrophages and dendritic cells, thus providing a novel interpretation to its immunosuppressive properties. 1,25(OH)2D3 significantly inhibits mRNA expression for both IL-12 p35 and p40 subunits acting at the transcriptional level. The effect of 1,25(OH)2D3 on p40 promoter activation was analyzed by cotransfecting monocytic RAW264.7 cells with p40 promoter/reporter constructs and expression vectors for vitamin D3 receptor (VDR) and/or retinoid X receptor (RXRalpha). We observed transcriptional repression of the p40 gene by 1,25(OH)2D3, which required coexpression of VDR with RXR and an intact VDR DNA-binding domain. The repressive effect maps to a region in the p40 promoter containing a binding site for NF-kappaB (p40-kappaB). Deletion of the p40-kappaB site abrogates part of the inhibitory effect on the p40 promoter, confirming the functional relevance of this site. Activation of monocytic THP-1 cells in the presence of 1,25(OH)2D3 results in reduced binding to the p40-kappaB site. Thus, 1,25(OH)2D3 may negatively regulate IL-12 production by downregulation of NF-kappaB activation and binding to the p40-kappaB sequence.

Measuring molecular frequencies in the 1-10 µm range at 11-digits accuracy.

High-resolution spectroscopy in the 1-10 µm region has never been fully tackled for the lack of widely-tunable and practical light sources. Indeed, all solutions proposed thus far suffer from at least one of three issues: they are feasible only in a narrow spectral range; the power available for spectroscopy is limited; the frequency accuracy is poor. Here, we present a setup for high-resolution spectroscopy, whose approach can be applied in the whole 1-10 µm range. It combines the power of quantum cascade lasers (QCLs) and the accuracy achievable by difference frequency generation using an orientation patterned GaP crystal. The frequency is measured against a primary frequency standard using the Italian metrological fibre link network. We demonstrate the performance of the setup by measuring a vibrational transition in a highly-excited metastable state of CO around 6 µm with 11 digits of precision.

First description of the soft part anatomy of Ilyocypris ramirezi Cusminsky & Whatley (Crustacea, Ostracoda) from Argentina, South America.

The anatomy of the soft parts of Ilyocypris ramirezi Cusminsky & Whatley, 1996 is described and illustrated for the first time, based on findings of this species from water bodies in the shallow areas of the Llancanelo basin, south-west of Mendoza Province, Argentina. This species is common in Quaternary and extant environments of the Pampa and Patagonian regions. Its distribution is now extending in Argentina to the Central-West area, locally named "Cuyo region". Ilyocypris ramirezi is a good environmental indicator and constitutes a useful tool in paleolimnological studies.

The amino acid sequence of the insulin receptor is normal in an insulin-resistant Pima Indian.

Insulin resistance is an early predictor of development of noninsulin-dependent diabetes mellitus (NIDDM) in Pima Indians, a population with the highest reported prevalence of NIDDM. The insulin receptor plays a central role in mediating insulin action, and previous studies have demonstrated that mutations in the insulin receptor gene may cause insulin resistance. Therefore, we have cloned the insulin receptor cDNA from an insulin-resistant Pima Indian to determine if there is a mutation in the patient's insulin receptor gene. We obtained nine cDNA clones spanning exons 4-10 and 12-22 of the patient's insulin receptor gene. Polymorphisms in the nucleotide sequences for codons 523 (Ala), 1058 (His), and 1062 (Leu) provided useful markers to differentiate the patient's two alleles of the insulin receptor gene. These substitutions were silent, in that they did not alter the predicted amino acid sequence. The sequence of exons 1-3 and 11 was determined directly from genomic DNA that had been amplified using the polymerase chain reaction catalyzed by Taq DNA polymerase. Other investigators have reported defects in insulin binding and insulin receptor tyrosine kinase activity in diabetic Pima Indians. However, we did not detect any mutations in this patient's insulin receptor gene. Thus, these observations are consistent with the interpretation that the defects in insulin receptor function are acquired rather than derived from defects in the primary structure of the receptor.

The SHIP phosphatase becomes associated with Fc gammaRIIB1 and is tyrosine phosphorylated during 'negative' signaling.

Immune-complex mediated co-ligation of antigen and Fc receptors on B-cells leads to abortive antigen receptor (BCR) signaling and provides a mechanism for feedback regulation of the immune response. A phosphotyrosine-containing 13 amino acid sequence (ITIM) found in the FcgammaRIIB1 cytoplasmic tail mediates this inhibition and specifically associates with the phosphotyrosine phosphatase SHP1. In vitro binding studies demonstrate that the phosphorylated ITIM binds unidentified proteins of 70 and 160 kD in addition to SHP1. Here we report the identification of p70 as SHP2 and p160 as the SH2 containing phosphatidylinositol 3,4,5-trisphosphate 5-phosphatase SHIP. SHIP is inducibly tyrosine phosphorylated following BCR-FcgammaRIIB1 co-ligation. Further, we observe SHIP association with tyrosine phosphorylated FcgammaRIIB1 in intact cells following BCR-FcgammaRIIB1 co-ligation. To a much lesser but significant degree, tyrosine phosphorylation of SHIP is also observed upon BCR ligation. These observations suggest that SHIP may play an important role in FcgammaRIIB1 dependent and independent regulation of BCR signaling.

Type I interferons and the Th1/Th2 paradigm.

During the past year significant advances have been made in our understanding of the factors contributing to the differentiation of CD4+ T helper cell subsets. These have been driven, in part, by the realization that cytokines from the innate immune response, such as interleukin-12 (IL-12) and interferons (IFNs), play a critical role in T cell subset differentiation. This review covers some of the most recent data concerning the divergent role that IFNs have in the differentiation of human versus mouse T helper cell subsets. In this review we discuss the molecular basis for the specie-specific effect of type I IFN on the selective induction of Th1 type immune responses. Furthermore, since IFN-beta is used in the treatment of multiple sclerosis (MS) we discuss the potential effects of such treatment and the value of the Th1/Th2 paradigm in MS.

Clinical pharmacology of ponesimod, a selective S1P1 receptor modulator, after uptitration to supratherapeutic doses in healthy subjects.

PURPOSE: The aim of this study was to assess in healthy subjects the safety, tolerability, pharmacokinetics, and pharmacodynamics of ponesimod, an oral selective sphingosine-1-phosphate receptor 1 (S1P1) modulator in development for multiple sclerosis, by using an uptitration scheme up to supratherapeutic doses. METHODS: This was a double-blind, placebo-controlled, randomised, parallel group, uptitration study. Male and female subjects received ascending oral doses of ponesimod (n=12) or placebo (n=4) once daily for 3 days at each dose level (10-20-40-60-80-100mg). RESULTS: The most frequent adverse events were chest discomfort, headache, dizziness, dyspnoea, abdominal pain, and night sweats. Chest discomfort and dyspnoea were considered dose-limiting. A transient decrease in heart rate was observed following the first 10-mg ponesimod dose (maximum mean decrease of 9 beats per minute (bpm) (placebo: 2 bpm)). After uptitration, effects on heart rate were indistinguishable from placebo. A dose-dependent effect on pulmonary function tests was observed and reached a plateau with 60-80 mg ponesimod (maximum mean decrease from baseline of 1.24l (-30.5%) in forced expiratory volume in 1s). A plateau in mean lymphocyte count reduction of approximately 70% from baseline was reached at the 40 mg dose level. Observed effects were fully reversible within 10days after treatment discontinuation. No relevant sex differences were observed. CONCLUSIONS: At supratherapeutic doses, symptoms of chest discomfort and dyspnoea were dose-limiting. An uptitration dosing scheme is to be preferred in clinical studies in patients in order to limit effects of ponesimod on heart rate and atrioventricular (AV) conduction.

Accurate modeling of a DOI capable small animal PET scanner using GATE.

In this work we developed a Monte Carlo (MC) model of the Sedecal Argus pre-clinical PET scanner, using GATE (Geant4 Application for Tomographic Emission). This is a dual-ring scanner which features DOI compensation by means of two layers of detector crystals (LYSO and GSO). Geometry of detectors and sources, pulses readout and selection of coincidence events were modeled with GATE, while a separate code was developed in order to emulate the processing of digitized data (for example, customized time windows and data flow saturation), the final binning of the lines of response and to reproduce the data output format of the scanner's acquisition software. Validation of the model was performed by modeling several phantoms used in experimental measurements, in order to compare the results of the simulations. Spatial resolution, sensitivity, scatter fraction, count rates and NECR were tested. Moreover, the NEMA NU-4 phantom was modeled in order to check for the image quality yielded by the model. Noise, contrast of cold and hot regions and recovery coefficient were calculated and compared using images of the NEMA phantom acquired with our scanner. The energy spectrum of coincidence events due to the small amount of (176)Lu in LYSO crystals, which was suitably included in our model, was also compared with experimental measurements. Spatial resolution, sensitivity and scatter fraction showed an agreement within 7%. Comparison of the count rates curves resulted satisfactory, being the values within the uncertainties, in the range of activities practically used in research scans. Analysis of the NEMA phantom images also showed a good agreement between simulated and acquired data, within 9% for all the tested parameters. This work shows that basic MC modeling of this kind of system is possible using GATE as a base platform; extension through suitably written customized code allows for an adequate level of accuracy in the results. Our careful validation against experimental data confirms that the developed simulation setup is a useful tool for a wide range of research applications.