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Certain regions on the surfaces of developing pollen grains exhibit very limited deposition of pollen wall exine. These regions give rise to pollen apertures, which are highly diverse in their patterns and specific for individual species. Arabidopsis thaliana pollen develops three equidistant longitudinal apertures. The precision of aperture formation suggests that, to create them, pollen employs robust mechanisms that generate distinct cellular domains. To identify players involved in this mechanism, we screened natural Arabidopsis accessions and discovered one accession, Martuba, whose apertures form abnormally due to the disruption of the protein kinase D6PKL3. During pollen development, D6PKL3 accumulates at the three plasma membrane domains underlying future aperture sites. Both D6PKL3 localization and aperture formation require kinase activity. Proper D6PKL3 localization is also dependent on a polybasic motif for phosphoinositide interactions, and we identified two phosphoinositides that are specifically enriched at the future aperture sites. The other known aperture factor, INAPERTURATE POLLEN1, fails to aggregate at the aperture sites in d6pkl3 mutants, changes its localization when D6PKL3 is mislocalized, and, in turn, affects D6PKL3 localization. The discovery of aperture factors provides important insights into the mechanisms cells utilize to generate distinct membrane domains, develop cell polarity, and pattern their surfaces. [ABSTRACT FROM AUTHOR]

Pollen presents a powerful model for studying mechanisms of precise formation and deposition of extracellular structures. Deposition of the pollen wall exine leads to the generation of species-specific patterns on pollen surface. In most species, exine does not develop uniformly across the pollen surface, resulting in the formation of apertures–openings in the exine that are species-specific in number, morphology and location. A long time ago, it was proposed that number and positions of apertures might be determined by the geometry of tetrads of microspores–the precursors of pollen grains arising via meiotic cytokinesis, and by the number of last-contact points between sister microspores. We have tested this model by characterizing Arabidopsis mutants with ectopic apertures and/or abnormal geometry of meiotic products. Here we demonstrate that contact points per se do not act as aperture number determinants and that a correct geometric conformation of a tetrad is neither necessary nor sufficient to generate a correct number of apertures. A mechanism sensitive to pollen ploidy, however, is very important for aperture number and positions and for guiding the aperture factor INP1 to future aperture sites. In the mutants with ectopic apertures, the number and positions of INP1 localization sites change depending on ploidy or ploidy-related cell size and not on INP1 levels, suggesting that sites for aperture formation are specified before INP1 is brought to them. [ABSTRACT FROM AUTHOR]

Abstract: Reproductive success of angiosperms relies on the precise development of the gynoecium and the anther, because their primary function is to bear and to nurture the embryo sac/female gametophyte and pollen, in which the egg and sperm cells, respectively, are generated. It has been known that the GRF-INTERACTING FACTOR (GIF) transcription co-activator family of Arabidopsis thaliana (Arabidopsis) consists of three members and acts as a positive regulator of cell proliferation. Here, we demonstrate that GIF proteins also play an essential role in development of reproductive organs and generation of the gamete cells. The gif1 gif2 gif3 triple mutant, but not the single or double mutants, failed to establish normal carpel margin meristem (CMM) and its derivative tissues, such as the ovule and the septum, resulting in a split gynoecium and no observable embryo sac. The gif triple mutant also displayed severe structural and functional defects in the anther, producing neither microsporangium nor pollen grains. Therefore, we propose that the GIF family of Arabidopsis is a novel and essential component required for the cell specification maintenance during reproductive organ development and, ultimately, for the reproductive competence. [Copyright &y& Elsevier]

In an effort to elucidate biological functions of transcription factors of Brassica rapa L. (ssp. pekinensis), an NGATHA homolog, BrNGA1, that belongs to the B3-type transcription factor superfamily was identified and expressed in Arabidopsis thaliana under the control of the cauliflower mosaic virus (CaMV) 35S promoter. Arabidopsis plants overexpressing BrNGA1, named BrNGA1ox, displayed markedly reduced organ growth compared with the wild type: lateral organs, such as leaves, flowers and cotyledons, were small and distinctively narrow, and their root growth was also severely retarded. Reduced sizes of BrNGA1ox organs were mainly due to reduction in cell numbers. Kinematic analysis of leaf growth revealed that both the rate and duration of cell proliferation declined during organogenesis, which was consistent with the reduced expression of cyclin genes. Reduction in organ growth was strongly correlated with the small size of meristematic cell pools in the shoot and root meristems. Taken together, these data indicate that BrNGA1 acts as a negative regulator of cell proliferation and may do so, in part, by regulating the size of the meristematic cell pool. [ABSTRACT FROM PUBLISHER]

Pulmonary delivery provides the most promising non-parenteral route of insulin administration. Insulin was used as a model protein to demonstrate the feasibility of using protein crystals for the pulmonary delivery of a sustained-release protein drug formulation. Insulin microcrystals with a mean diameter of 3 μm were prepared using a seed zone method. The yield of crystallization was very high (95.8±0.97%), and the microcrystals were recovered with high efficiency (>98%) by centrifugation. Morphological examination using scanning electron microphotography showed the microcrystals to be of a homogeneous rhombohedral shape, with some rhombus forms, without aggregates. After the administration of 32 U/kg of the microcrystal suspension to STZ-induced diabetic SD rats by intratracheal instillation, the blood glucose levels were reduced and hypoglycemia was prolonged over 13 h, as compared to the insulin solution. The percent minimum reductions of the blood glucose concentration (% MRBG) produced by the microcrystal suspension and insulin solution reached 36.5 and 37.2%, respectively, of the initial level, and the percent total reductions in blood glucose (% TRBG13h) were 34.4 and 25.0%, respectively. In the case of inhalation using a sieve-type ultrasonic nebulizer, the % MRBG produced by the microcrystal suspension and insulin solution were 21.7 and 26.3%, respectively, of the initial level, and the % TRBG13h were 66.7 and 58.4%, respectively. However, the hypoglycemic effects of the microcrystal suspension were prolonged over 7 h, which compares favorably with the insulin solution (P<0.05 by unpaired t-test). These results could be attributed to the sustained-release of insulin from the microcrystals, which were deposited widely throughout the entire lung. [Copyright &y& Elsevier]

A low lymphocyte count puts immune‐compromised patients at risk of mortality. hIL‐7‐hyFc is a homodimeric interleukin‐7 (IL‐7), a potent T‐cell amplifier, fused to the hybridizing IgD/IgG4 immunoglobulin domain. We performed a randomized, double‐blind, placebo‐controlled, dose‐escalation, phase I study to assess the pharmacokinetic, pharmacodynamic, safety, tolerability, and immunogenicity profiles of hIL‐7‐hyFc administered s.c. and i.m. to healthy volunteers. Thirty subjects randomly received hIL‐7‐hyFc or its matching placebo in an 8:2 ratio at 20, 60 μg/kg s.c., or 60 μg/kg i.m. The hIL‐7‐hyFc was slowly absorbed and its terminal half‐life was 63.26 hours after i.m. administration. The hIL‐7‐hyFc increased absolute lymphocyte count, mostly in T‐cells, which peaked 3 weeks after administration and then lasted for several additional weeks. The hIL‐7‐hyFc was well‐tolerated after a single s.c. and i.m. administration. Injection site reaction was the most common treatment‐emergent adverse event, which resolved spontaneously without treatment. The hIL‐7‐hyFc can be developed into a beneficial treatment option for patients with compromised T‐cell immunity. This trial was registered at www.clinicaltrials.gov as #NCT02860715. [ABSTRACT FROM AUTHOR]

Objectives: Emerging oncotherapeutic strategies require the induction of an immunostimulatory tumor microenvironment (TME) containing numerous tumor‐reactive CD8+ T cells. Interleukin‐7 (IL‐7), a T‐cell homeostatic cytokine, induces an antitumor response; however, the detailed mechanisms underlying the contributions of the IL‐7 to TME remain unclear. Here, we aimed to investigate the mechanism underlying the induction of antitumor response by hybrid Fc‐fused long‐acting recombinant human IL‐7 (rhIL‐7‐hyFc) through regulation of both adaptive and innate immune cells in the TME. Methods: We evaluated rhIL‐7‐hyFc‐mediated antitumor responses in murine syngeneic tumor models. We analysed the cellular and molecular features of tumor‐infiltrating lymphocytes (TILs) and changes in the TME after rhIL‐7‐hyFc treatment. Furthermore, we evaluated the antitumor efficacy of rhIL‐7‐hyFc combined with chemotherapy and checkpoint inhibitors (CPIs). Results: Systemic delivery of rhIL‐7‐hyFc induced significant therapeutic benefits by expanding CD8+ T cells with enhanced tumor tropism. In tumors, rhIL‐7‐hyFc increased both tumor‐reactive and bystander CD8+ TILs, all of which displayed enhanced effector functions but less exhausted phenotypes. Moreover, rhIL‐7‐hyFc suppressed the generation of immunosuppressive myeloid cells in the bone marrow of tumor‐bearing mice, resulting in the immunostimulatory TME. Combination therapy with chemotherapy and CPIs, rhIL‐7‐hyFc elicited a strong antitumor response and even under a T lymphopenic condition by restoring CD8+ T cells. When combined with chemotherapy and CPIs, rhIL‐7‐hyFc administration enhanced antitumor response under intact andlymphopenic conditions by restoring CD8+ T cells. Conclusion: Taken together, these data demonstrate that rhIL‐7‐hyFc induces antitumor responses by generating T‐cell‐inflamed TME and provide a preclinical proof of concept of immunotherapy with rhIL‐7‐hyFc to enhance therapeutic responses in the clinic. [ABSTRACT FROM AUTHOR]

Studying the performance limiting parameters of the Schottky device is an urgent issue, which are addressed herein by thermally stable silver nanowire (AgNW) embedded metal oxide/p-Si Schottky device. Temperature and bias dependent junction interfacial properties of AgNW-ITO/Si Schottky photoelectric device are reported. The current−voltage−temperature ( I − V − T ), capacitance-voltage-temperature ( C − V − T ) and impedance analysis have been carried out in the high-temperature region. The ideality factor and barrier height of Schottky junction are assessed using I − V − T characteristics and thermionic emission, to reveal the decrease of ideality factor and increase of barrier height by the increasing of temperature. The extracted values of laterally homogeneous Schottky (ϕ b ) and ideality factor ( n) are approximately 0.73 eV and 1.58, respectively. Series resistance (R s ) assessed using Cheung's method and found that it decreases with the increase of temperature. A linear response of R s of AgNW-ITO/Si Schottky junction is observed with respect to change in forward bias, i.e. dR S /dV from 0 to 0.7 V is in the range of 36.12–36.43 Ω with a rate of 1.44 Ω/V. Impedance spectroscopy is used to study the effect of bias voltage and temperature on intrinsic Schottky properties which are responsible for photoconversion efficiency. These systematic analyses are useful for the AgNWs-embedding Si solar cells or photoelectrochemical cells. [ABSTRACT FROM AUTHOR]

The precursors for the formation of thermoelectric skutterudite CoSb 3 nanoparticles are predicted by thermodynamic modeling of the complex chemical species. Based on the results, equimolar mixture of CoC 2 O 4 ·2H 2 O and Sb(C 2 O 4 )OH are successively co-precipitated under controlled conditions of pH = 2.7 and concentration of reactants. The as synthesized powder was decomposed at 350 °C to remove the organic molecules and further reduced to CoSb 3 phase by heating at 530 °C under hydrogen flow. The obtained powder was consolidated by spark plasma sintering (SPS). CoSb 3 prepared by controlled chemical co-precipitation has p-type behavior with a positive sign of the Seebeck coefficient. TE transport properties were measured, which revealed that the Seebeck coefficient increased 2.5 times with increasing the temperature and it is lower than the ball milled CoSb 3 . Thermal conductivity of sintered CoSb 3 at 773 K starts from 0.06 W/cm K at room temperature and decreases to 0.04 W/cm K at 700 K, which is lower than the bulk counterpart. The ZT of coprecipitated CoSb 3 and SPS consolidated at 773 K shows 2 times higher than the ball milled one. [ABSTRACT FROM AUTHOR]

Background Sézary syndrome (SS) is a highly-morbid T cell leukemic lymphoma with no widely-effective treatments and few preclinical models. SS T cells typically lose CD7 but maintain ubiquitous high CD2 expression. Thus, we generated CD2 - and TRAC -deleted anti-CD2 universal CARTs (UCART2) and multiple SS xenograft models (PDXs) as preclinical UCART2 testing platforms. We further tested a stable homodimeric interleukin-7 molecule, the long-acting form of recombinant human interleukin-7 fused with hybrid Fc (rhIL-7-hyFc, NT-I7), to potentiate UCART2 killing of an SS xenograft in vivo. Methods To generate SS PDX models, we injected NSG-SGM3 mice with ∼2 × 106 mononuclear cells derived from SS patients. We immunophenotyped SS patient blood and PDX engraftment with two 21-color flow cytometry panels assessing major immune subsets, CTCL, and exhaustion markers. To generate UCART2s, we CRISPR/Cas9 multiplexed edited CD2 and TRAC from CART2. For initial testing, we injected NSG mice with 5 × 105 cells from a human Sézary cell line (HH) on day -4 from UCART2 treatment, followed by NT-I7 (10mg/kg SC) on days +1, +15 and +29. Results SS patient blood showed specific defects in monocyte, monocytic dendritic cell, and natural killer cell differentiation, increased skewing toward granulocytes and non-classical CD16+ monocytes (p<0.01, SS vs. normal PBMCs), and loss of effector memory CD4 cells (8% vs 34%, p<0.001, SS vs. normal PBMCs). SS cells were CD3+CD4+CD2+CD5+CD8- with variable CD7 loss and PD1 gain. Four of six unique human SS samples injected in NSG-SGM3 mice engrafted within ∼6 weeks with no signs of xenogeneic GVHD. Following engraftment, SS cells showed near ubiquitous PD1 expression (>90% vs ∼20%, p<0.001, SS vs. normal PBMCs), CD7 loss, and increased CD30 and CD26 expression. Immunohistochemistry further revealed atypical CD3+CD4+CD8-CD7- lymphocytes lining the dermo-epidermal junction. UCART2-treated HHCBR-GFP mice showed dramatically reduced tumor burden as compared to control UCART19-treated HHCBR-GFP mice (BLI; 10^7 vs. 10^11 photon flux/s at 3 weeks, p<0.0001, UCART2 vs. UCART19). Moreover, UCART2-treated HHCBR-GFP mice survived ∼40 days as compared to ∼21 days in the UCART19 group. Remarkably, UCART2-treated HHCBR-GFP mice receiving NT-I7 showed virtually no tumor burden (maximum 106 photon flux/s vs. 1010 photon flux/s, UCART2+NT-I7 vs. UCART2 only groups) with 100% of UCART2- and NT-I7-treated HHCBR-GFP mice surviving beyond 70 days. Discussion We describe the generation of physiologically-relevant SS preclinical models and the highly effective anti-tumor activity of UCART2- plus NT-I7-mediated killing of SS cells in vivo using an NSG xenograft model. [ABSTRACT FROM AUTHOR]

Background CART therapy is revolutionizing modern cancer therapy, with two anti-CD19 CARTs FDA-approved for relapsed/refractory B cell lymphoma/leukemia and many other CARTs for solid and liquid tumors currently undergoing clinical trials. However, in current clinical practice, suboptimal CART persistence and tumor killing permit tumor cell escape and, ultimately, disease relapse. Reasoning that a pro-lymphoid growth factor could promote CART efficacy, we supplemented UCART infusion with subcutaneous injections of the long-acting form of recombinant human interleukin-7 fused with hybrid Fc (rhIL-7-hyFc, NT-I7) in vivo using a CD19+ lymphoma xenograft model. Methods To create anti-CD19 universal CART (UCART19), we activated human T cells on CD3/CD28 beads, electroporated the T cells with Cas9 mRNA and a TRAC gRNA, and virally transduced an anti-CD19 scFv 3rd generation CAR containing a peptidase 2A-cleaved human CD34 construct for both purification and tracking in vivo. Residual TRAC+ cells were depleted using magnetic selection. For xenograft tumor modeling in vivo, we injected NSG mice with 5 × 105 RamosCBR-GFP cells four days prior to UCART19 (2 × 106 cells). Mice were treated with NT-I7 (10mg/kg SC) on days +1, +15 and +29 post UCART19 infusion. Results RamosCBR-GFP mice receiving NT-I7 without UCART19 (NT-I7 only group) survived marginally longer (24 day med survival) than mice receiving RamosCBR-GFP cells alone (No tx group) (21 day medium survival, p=0.018, NT-I7 only vs. No Tx). While RamosCBR-GFP mice treated with UCART19 alone (UCART19 group) survived 33 days, 100% of RamosCBR-GFP mice treated with UCART19 and NT-I7 (UCART19+NT-I7 group) were alive at 80 days (Fig 1a), with no mouse showing signs of xenogeneic GVHD (p<0.0001, UCART19+NT-I7 vs. UCART19). At three weeks post UCART19 infusion, bioluminescent imaging (BLI) revealed minimal tumor signal in UCART19+NT-I7 treated mice (108 vs. 1010 photon flux/s, p<0.05, UCART19+NT-I7 vs. UCART19) and near-undetectable photon flux/s at four weeks (107 vs 1011 photon flux/s, p<0.0001, UCART19+NT-I7 vs. UCART19). Quantitative 17-parameter flow cytometric analyses of the blood, bone marrow, and spleens revealed an up to ∼8000-fold increase in UCART19 cells in NT-I7-treated mice four weeks post UCART19 infusion (Fig 1a). These UCART19 cells demonstrated a predominantly effector and effector memory phenotype. Discussion Here, we demonstrate that a pharmacological grade long-acting interleukin-7 agonist can potentiate adoptive cellular therapies. Specifically, NT-I7 can dramatically enhance gene modified T cell proliferation, persistence and tumor killing in vivo, resulting in enhanced survival. Unlike genetic strategies to potentiate IL-7 signaling in CART, NT-I7 provides a tunable clinic-ready adjuvant for reversing suboptimal CART activity in vivo. [ABSTRACT FROM AUTHOR]

The immune system plays an important role in maintaining body homeostasis. Recent studies on the immune-enhancing effects of ginseng saponins have revealed more diverse mechanisms of action. Maillard reaction that occurs during the manufacturing processes of red ginseng produces a large amount of Amadori rearrangement compounds (ARCs), such as arginyl-fructose (AF). The antioxidant and anti-hyperglycemic effects of AF have been reported. However, the possible immune enhancing effects of non-saponin ginseng compounds, such as AF, have not been investigated. In this study the effects of AF and AF-enriched natural product (Ginofos, GF) on proliferation of normal mouse splenocytes were evaluated in vitro and male BALB/c mice models. The proliferation of splenocytes treated with mitogens (concanavalin A, lipopolysaccharide) were further increased by addition of AF (p < 0.01) or GF (p < 0.01), in a dose dependent manner. After the 10 days of oral administration of compounds, changes in weights of spleen and thymus, serum immunoglobulin, and expression of cytokines were measured as biomarkers of immune-enhancing potential in male BALB/c mice model. The AF or GF treated groups had higher weights of the thymus (0.94 ± 0.25 and 0.86 ± 0.18, p < 0.05, respectively) than that of cyclophosphamide treated group (0.59 ± 0.18). This result indicates that AF or AF-enriched extract (GF) increased humoral immunity against CY-induced immunosuppression. In addition, immunoglobulin contents and expression of cytokines including IgM (p < 0.01), IgG (p < 0.05), IL-2 (p < 0.01), IL-4 (p < 0.01), IL-6 (p < 0.01), and IFN-γ (p < 0.05) were also significantly increased by supplementation of AF or GF. These results indicate that AF has immune enhancing effects by activation of adaptive immunity via increase of expression of immunoglobulins and cytokines such as IgM, IgG, IL-2, IL-4, IL-6 and thereby proliferating the weight of thymus. Our findings provide a pharmacological rationale for AF-enriched natural products such as ginseng and red ginseng that can possibly have immune-enhancement potential and should be further evaluated. [ABSTRACT FROM AUTHOR]

Novel engineered T cells containing chimeric antigen receptors (CAR-T cells) that combine the benefits of antigen recognition and T cell response have been developed, and their effect in the anti-tumor immunotherapy of patients with relapsed/refractory leukemia has been dramatic. Thus, CAR-T cell immunotherapy is rapidly emerging as a new therapy. However, it has limitations that prevent consistency in therapeutic effects in solid tumors, which accounts for over 90% of all cancer patients. Here, we review the literature regarding various obstacles to CAR-T cell immunotherapy for solid tumors, including those that cause CAR-T cell dysfunction in the immunosuppressive tumor microenvironment, such as reactive oxygen species, pH, O2, immunosuppressive cells, cytokines, and metabolites, as well as those that impair cell trafficking into the tumor microenvironment. Next-generation CAR-T cell therapy is currently undergoing clinical trials to overcome these challenges. Therefore, novel approaches to address the challenges faced by CAR-T cell immunotherapy in solid tumors are also discussed here. [ABSTRACT FROM AUTHOR]

A new type of ultrasonic rotary motor that can replace existing ultrasonic motors for driving camera zoom lenses was proposed and investigated experimentally. The proposed motor composed of four roof-shaped stators. On the upper surfaces of each roof-shaped stator, two rectangular ceramics were attached. When two AC voltages with a 90° phase difference were applied to the ceramics, elliptical displacements occurred at the tip line of the actuator. By combining these elliptical displacements of the four stators, rotational motion of the stator could be achieved. If a circular disk rotor was positioned on the stator, vibrations of the stator were transferred through the four contact lines between the stator and the rotor. Preload of the motor could be controlled by the jointing bolt, which went through the center of the disk rotor and the ball bearings. The finite element analysis (FEA) simulation program ANSYS was used to analyze the elliptical displacement characteristics in relation to changes in the size of the elastic body, piezoelectric ceramics, friction tips, frequency, and applied voltages. A prototype motor was fabricated, and its driving characteristics such as impedances, vibration amplitudes, rotational speed, and torque were measured. [ABSTRACT FROM AUTHOR]