Naïve B cells with low differentiation improve the immune reconstitution of HIV-infected patients.

Incomplete immune reconstitution happens in some HIV-infected patients who have achieved persistent viral suppression under antiretroviral therapy (ART). We performed single-cell RNA sequencing for peripheral blood mononuclear cells to analyze B cell receptor (BCR) repertoire and B cell subtypes in health controls (non-HIV-infected, HCs), HIV-infected immunological responders (IRs), and immunological nonresponders (INRs). We found that the dominant usage of IGHV gene segments of naïve B cells and memory B cells were IGHV3 and IGHV4, and the diversity of BCR repertoire was decreased in INRs. Differentiation trajectory analysis showed that the low differentiation of naïve B cells was related to satisfactory immune status. The cell cycle of B cells with immune-specific genes of IgD+ B cells was degraded in INRs, which was mediated by the anaphase-promoting complex/cyclosome pathway in the phase of G2/M checkpoints. These findings provide significant insights to understand the function of B cell-mediated immune response in immune reconstitution after HIV infection.

Flavonoid glycosides isolated from Epimedium brevicornum and their estrogen biosynthesis-promoting effects.

Epimedium brevicornum Maxim has a long history of use in the treatment of estrogen deficiency-related diseases. However, the chemical constituents and mechanism of action of this medicinal plant are not fully understood. In the present study, we isolated four new isoprenylated flavonoid glycosides, as well as 16 known flavonoids (13 isoprenylated flavonoids), from this plant. The chemical structures of the new flavonoid glycosides were elucidated by extensive spectroscopic analysis. The new compounds 1-4 were potent promoters of estrogen biosynthesis in human ovarian granulosa-like KGN cells. ZW1, an isoprenylated flavonoid analogue and a specific inhibitor of phosphodiesterase 5 (PDE5), was synthesized and used to explore the mechanism of the isoprenylated analogues on estrogen biosynthesis. ZW1 treatment increased estrogen production by upregulation of aromatase mRNA and protein expression. ZW1 increased the phosphorylation of cAMP response element-binding protein (CREB). Further study showed that the inhibition of PDE5 by ZW1 increased estrogen biosynthesis partly through suppression of phosphodiesterase 3 (PDE3). Our results suggested that the isoprenylated flavonoids from E. brevicornum may produce beneficial health effects through the promotion of estrogen biosynthesis. PDE5 warrants further investigation as a new therapeutic target for estrogen biosynthesis in the prevention and treatment of estrogen-deficiency related diseases.

Nanoporous Gold Films Prepared by a Combination of Sputtering and Dealloying for Trace Detection of Benzo[a]pyrene Based on Surface Plasmon Resonance Spectroscopy.

A wavelength-interrogated surface plasmon resonance (SPR) sensor based on a nanoporous gold (NPG) film has been fabricated for the sensitive detection of trace quantities of benzo[a]pyrene (BaP) in water. The large-area uniform NPG film was prepared by a two-step process that includes sputtering deposition of a 60-nm-thick AuAg alloy film on a glass substrate and chemical dealloying of the alloy film in nitric acid. For SPR sensor applications, the NPG film plays the dual roles of analyte enrichment and supporting surface plasmon waves, which leads to sensitivity enhancement. In this work, the as-prepared NPG film was first modified with 1-dodecanethiol molecules to make the film hydrophobic so as to improve BaP enrichment from water via hydrophobic interactions. The SPR sensor with the hydrophobic NPG film enables one to detect BaP at concentrations as low as 1 nmol·L-1. In response to this concentration of BaP the sensor produced a resonance-wavelength shift of ΔλR = 2.22 nm. After the NPG film was functionalized with mouse monoclonal IgG1 that is the antibody against BaP, the sensor's sensitivity was further improved and the BaP detection limit decreased further down to 5 pmol·L-1 (the corresponding ΔλR = 1.77 nm). In contrast, the conventional SPR sensor with an antibody-functionalized dense gold film can give a response of merely ΔλR = 0.9 nm for 100 pmol·L-1 BaP.

Algorithmic Enhancement of Spectral Resolution of a LiNbO3 Waveguide-Based Miniature Fourier Transform Spectrometer.

In a recent report we demonstrated a miniature static Fourier transform spectrometer (FTS) that was implemented with a LiNbO3 (LN) waveguide electro-optic modulator (EOM) combined with the dispersion relation between its half-wave voltage and wavelength. The FTS was verified to be able to measure laser wavelength and for low-resolution spectroscopy. In this report, we successfully applied the resolution enhancement algorithm to the FTS, resulting in at least a three-fold increase in its spectral resolution without causing obvious distortion of the measured spectra. The algorithm method used is based on an autoregressive (AR) model, singular value decomposition (SVD), and forward-backward linear prediction (FBLP). The combination of these methods allows the FTS to remain a small size but to possess good spectral resolution, effectively mitigating the conflict between the small size and high resolution of the device. This study opens the way to development of high-resolution miniature FTS.

Fiber optofluidic biosensor for the label-free detection of DNA hybridization and methylation based on an in-line tunable mode coupler.

An optical fiber optofluidic biosensor for the detection of DNA hybridization and methylation has been proposed and experimentally demonstrated. An in-line fiber Michelson interferometer was formed in the photonic crystal fiber. A micrhole in the collapsed region, which combined the tunable mode coupler and optofluidic channel, was fabricated by using femtosecond laser micromachining. The mode field diameter of the guided light is changed with the refractive index in the optofluidic channel, which results in the tunable coupling ratio. Label-free detections of the DNA hybridization and methylation have been experimentally demonstrated. The probe single stranded DNA (ssDNA) was bound with the surface of the optofluidic channel through the Poly-l-lysine layer, and the hybridization between a short 22-mer probe ssDNA and a complementary target ssDNA was carried out and detected by interrogating the fringe visibility of the reflection spectrum. Then, the DNA methylation was also detected through the binding between the methylated DNA and the 5-methylcytosine (5-mC) monoclonal antibody. The experiments results demonstrate that the limit of detection of 5nM is achieved, establishing the tunable mode coupler as a sensitive and versatile biosensor. The sensitive optical fiber optofluidic biosensor possesses high specificity and low temperature cross-sensitivity.

A prototype stationary Fourier transform spectrometer for near-infrared absorption spectroscopy.

A prototype stationary Fourier transform spectrometer (FTS) was constructed with a fiber-coupled lithium niobate (LiNbO3) waveguide Mach-Zehnder interferometer (MZI) for the purpose of rapid on-site spectroscopy of biological and chemical measurands. The MZI contains push-pull electrodes for electro-optic modulation, and its interferogram as a plot of intensity against voltage was obtained by scanning the modulating voltage from -60 to +60 V in 50 ms. The power spectrum of input signal was retrieved by Fourier transform processing of the interferogram combined with the wavelength dispersion of half-wave voltage determined for the MZI used. The prototype FTS operates in the single-mode wavelength range from 1200 to 1700 nm and allows for reproducible spectroscopy. A linear concentration dependence of the absorbance at λmax = 1451 nm for water in ethanolic solution was obtained using the prototype FTS. The near-infrared spectroscopy of solid samples was also implemented, and the different spectra obtained with different materials evidenced the chemical recognition capability of the prototype FTS. To make this prototype FTS practically applicable, work on improving its spectral resolution by increasing the maximum optical path length difference is in progress.

Selective recruitment of host factors by HSV-1 replication centers.

Herpes simplex virus type 1 (HSV-1) enters productive infection after infecting epithelial cells, where it controls the host nucleus to make viral proteins, starts viral DNA synthesis and assembles infectious virions. In this process, replicating viral genomes are organized into replication centers to facilitate viral growth. HSV-1 is known to use host factors, including host chromatin and host transcription regulators, to transcribe its genes; however, the invading virus also encounters host defense and stress responses to inhibit viral growth. Recently, we found that HSV-1 replication centers recruit host factor CTCF but exclude γH2A.X. Thus, HSV-1 replication centers may selectively recruit cellular factors needed for viral growth, while excluding host factors that are deleterious for viral transcription or replication. Here we report that the viral replication centers selectively excluded modified histone H3, including heterochromatin mark H3K9me3, H3S10P and active chromatin mark H3K4me3, but not unmodified H3. We found a dynamic association between the viral replication centers and host RNA polymerase II. The centers also recruited components of the DNA damage response pathway, including 53BP1, BRCA1 and host antiviral protein SP100. Importantly, we found that ATM kinase was needed for the recruitment of CTCF to the viral centers. These results suggest that the HSV-1 replication centers took advantage of host signaling pathways to actively recruit or exclude host factors to benefit viral growth.

In situ study of self-assembled nanocomposite films by spectral SPR sensor.

Spectral surface plasmon resonance (SPR) sensor with a time-resolved charge-coupled device (CCD) detector is a powerful analytical tool for label-free detection of biomolecular interaction at the liquid/solid interface and for in situ study of molecular adsorption behavior. In this work, the layer-by-layer self-assembly processes for three nanocomposite films were monitored in real time using a broadband spectral SPR sensor with a large dynamic range. Kinetics studies suggest that cytochrome c (Cyt c) and deoxy ribonucleic acid (DNA) adsorptions obey the Langmuir-isotherm theory, while gold nanoparticle (GNP) adsorption follows the Diffusion-controlled model. Using poly(sodium 4-styrenesulfonate) (PSS) and poly(dimethyldiallylammonium chloride) (PDDA) as the positively charged agents, three kinds of multilayer films such as the PSS/Cyt c, GNP/Cyt c and PDDA/DNA binary nanocomposites were fabricated on the SPR chips by the electrostatic attraction based on self-assemble. The SPR response in terms of ΔλR was measured to linear increase with increasing the number of layers for a six-bilayer PSS/Cyt c nanocomposite film, indicating that every PSS/Cyt c layer has equal mass coverage. In contrast, the nonlinear dependences of ΔλR on the number of bilayers were observed for the GNP/Cyt c and PDDA/DNA nanocomposite multilayer films.

Waveguide-coupled directional Raman radiation for surface analysis.

Kretschmann-type waveguide structures, including Plasmon Waveguide (PW) and Resonant Mirror (RM), have been applied in interfacial Raman spectroscopy due to the following unique features: (1) unlike the classic surface enhanced Raman scattering (SERS) substrates made of either gold or silver, both PW and RM can be prepared using a large variety of inexpensive materials; (2) the field enhancement factors using these structures can be theoretically predicted and experimentally controlled, which enables us to manipulate the surface Raman sensitivity with high repeatability; (3) the use of transverse electric (TE) and transverse magnetic (TM) modes for Raman excitation allows us to evaluate the orientation of target molecules immobilized on the waveguide surface; (4) the unwanted impact of noble metals on the Raman fingerprints of target molecules, which is often observed for conventional SERS substrates, can be avoided upon the use of dielectric waveguides. In this paper, guided-mode-coupled directional Raman emission, which is an additional important feature of the waveguide Raman technique, was theoretically investigated based on the optical reciprocity theorem combined with the Fresnel equations. The simulation results indicate that the directional Raman emission from a dipole located within the field confinement and penetration depth of a guided mode depends on both the orientation of the dipole and its distance from the waveguide surface. Raman light from the TE-oriented dipoles is launched into the prism coupler at the TE-mode resonance angle and that from the non-TE-oriented dipoles propagates at the TM-mode resonance angle. The intensity of the guided-mode-excited Raman signal propagating at the mode resonance angle is proportional to the fourth power of the mode field (E(4)) at the depth of the dipole from the waveguide surface. This means that the guided-mode-excited and guided-mode-coupled directional Raman spectroscopy has a detection depth that is as small as a quarter of the evanescent-field penetration depth, indicating the excellent surface selectivity of this technique. The directional Raman emission also facilitates high-efficiency signal collection compared with conventional SERS. It is worth noting that Raman light from the dipoles confined in the core layer of a single-mode waveguide can be simultaneously coupled into both the guided mode and the substrate mode, especially the surface plasmon resonance (SPR) mode for PW.

Miniature Fourier transform spectrometer based on wavelength dependence of half-wave voltage of a LiNbO3 waveguide interferometer.

A simple and reliable spectrum-retrieval method was proposed for the development of miniature stationary Fourier transform (FT) spectrometers based on a LiNbO3 (LN) waveguide Mach-Zehnder interferometer (MZI) modulator. The method takes into account the wavelength dependence of the optical pathlength difference (OPD) and allows us to use a nonlinear voltage ramp to modulate the OPD. The method is based on the dispersion of the half-wave voltage, which was measured to be a monotonous polynomial function of the wavelength for the LN waveguide MZI used. With the measured dispersion of the half-wave voltage, the OPD, as a linear function of the modulating voltage, can be accurately determined at each wavelength in the near-infrared region in which the MZI used is a single-mode device. A prototype FT spectrometer was prepared using a LN waveguide MZI modulator based on the above method. The experimental results demonstrated that the spectrometer can be used for accurate determination of the laser wavelength and for liquid absorptiometry.

Wavelength-interrogated surface plasmon resonance sensor with mesoporous-silica-film-enhanced sensitivity to small molecules.

Sol-gel copolymer-templated mesoporous silica films with a thickness of 70 nm and interpore spacing of 4.34 nm were fabricated on gold layer covered glass substrates for application as a wavelength-interrogated surface plasmon resonance (SPR) sensor. The resonance wavelength (λ(R)) of the sensor with a solution sample was determined by absorptiometry at a given incident angle. A comparison between the experimental data obtained with the coated and uncoated SPR chips demonstrated that the mesoporous silica film effectively enhanced sensor response to individual adsorption of cysteamine molecules and lead(II) ions. An approximate proportional relationship between the resonance-wavelength shift of the sensor and the volume fraction of analyte molecules adsorbed in the mesoporous silica film was obtained by numerical simulation. Porosities of 0.865 and 0.785 for the two silica films used as well as the volume fractions of 0.048 and 0.116 for adsorbed lysozyme and cysteamine molecules were determined by fitting the simulation results to the experimental data. The adsorbed amount of cysteamine (∼0.5 nm) is equivalent to more than 16 full monolayers on the geometric surface of the mesoporous silica film used. In contrast, an equivalence of less than 2 full monolayers for adsorbed lysozyme molecules (3 nm × 3 nm × 4.5 nm) suggests that the mesoporous silica film has good size-selective adsorption capability due to its uniform pore size distribution. Cysteamine modification of the mesoporous silica film renders the SPR sensor able to detect lead(II) ions at concentrations as low as 1 nM.

Inhibitory effect of luteolin on estrogen biosynthesis in human ovarian granulosa cells by suppression of aromatase (CYP19).

Inhibition of aromatase, the key enzyme in estrogen biosynthesis, is an important strategy in the treatment of breast cancer. Several dietary flavonoids show aromatase inhibitory activity, but their tissue specificity and mechanism remain unclear. This study found that the dietary flavonoid luteolin potently inhibited estrogen biosynthesis in a dose- and time-dependent manner in KGN cells derived from human ovarian granulosa cells, the major source of estrogens in premenopausal women. Luteolin decreased aromatase mRNA and protein expression in KGN cells. Luteolin also promoted aromatase protein degradation and inhibited estrogen biosynthesis in aromatase-expressing HEK293A cells, but had no effect on recombinant expressed aromatase. Estrogen biosynthesis in KGN cells was inhibited with differing potencies by extracts of onion and bird chili and by four other dietary flavonoids: kaempferol, quercetin, myricetin, and isorhamnetin. The present study suggests that luteolin inhibits estrogen biosynthesis by decreasing aromatase expression and destabilizing aromatase protein, and it warrants further investigation as a potential treatment for estrogen-dependent cancers.

Biologically active steroids from the aerial parts of Vernonia anthelmintica Willd.

A new steroid, vernoanthelsterone A (1), and five known steroids were isolated from the aerial parts of Vernonia anthelmintica Willd. Compound 1 possesses a Δ(8(14))-15-one moiety. To our best knowledge, few steroids with this moiety have been reported before. Compounds 1-6 were tested for their antibacterial activities and their effects on estrogen biosynthesis in human ovarian granulosa-like cells (KGN cells). Compound 2 showed the ability to promote estrogen biosynthesis with EC(50) of 56.95 µg/mL and also exhibited the antibacterial activities against Bacillus cereus, Staphyloccocus aureus, Bacillus subtilis and Escherichia coli with MICs ranging from 3.15 to 15.5 µg/mL. The structures of 1-6 were determined on the basis of IR, MS, 1D and 2D NMR.

Kinetics of competitive adsorption of ß-casein and methylene blue on hydrophilic glass.

The competitive adsorption of methylene blue (MB) and ß-casein on hydrophilic glass from an aqueous mixed solution was directly detected at the solution pH smaller than the protein isoelectric point (pI) by means of the waveguide-based broadband time-resolved evanescent wave absorption spectroscopy. The competitive adsorption causes the MB coverage to exponentially decrease with time from its peak value and prevents MB aggregation at the interface. The kinetic equation for the competitive adsorption of binary adsorbates was theoretically deduced based on the Langmuir model, and was used for creating the best fit to the experimental data. In the case of a fixed concentration of MB in the mixed solution, the best-fit parameter τ(-1) increases with the protein concentration at a specific pH and decreases with the solution pH at a given concentration of protein. The findings suggest that the ß-casein concentration in sub-µM level can be rapidly determined by the time-resolved waveguide absorptiometry based on the competitive adsorption of MB and protein.

Determination of surface protein coverage by composite waveguide based polarimetric interferometry.

Deposition of a tapered thin film of Ta(2)O(5) onto a single-mode, polarization-insensitive slab glass waveguide enables the resultant structure to serve as a simple, inexpensive yet highly sensitive polarimetric interferometer for trace, even ultra-trace, detection of chemical and biochemical analytes. By comparing the measured refractive-index sensitivity with that simulated based on a four-layer homogeneous waveguide, the equivalent thickness for the tapered layer of Ta(2)O(5) and the sensitivity of the sensor to adlayer thickness were determined. Responses of the sensor to unspecific adsorption of bovine serum albumin (BSA) and to surface antibody-antigen interaction were investigated in situ and the corresponding surface coverages were obtained with the adlayer-thickness sensitivity. The interferometer sensor shows good long-term stability and its phase drift is lower than π over 10 h.