The dendritic cell lineage: ontogeny and function of dendritic cells and their subsets in the steady state and the inflamed setting.

Dendritic cells (DCs) form a remarkable cellular network that shapes adaptive immune responses according to peripheral cues. After four decades of research, we now know that DCs arise from a hematopoietic lineage distinct from other leukocytes, establishing the DC system as a unique hematopoietic branch. Recent work has also established that tissue DCs consist of developmentally and functionally distinct subsets that differentially regulate T lymphocyte function. This review discusses major advances in our understanding of the regulation of DC lineage commitment, differentiation, diversification, and function in situ.

The Helix-Loop-Helix Protein ID2 Governs NK Cell Fate by Tuning Their Sensitivity to Interleukin-15.

The inhibitor of DNA binding 2 (Id2) is essential for natural killer (NK) cell development with its canonical role being to antagonize E-protein function and alternate lineage fate. Here we have identified a key role for Id2 in regulating interleukin-15 (IL-15) receptor signaling and homeostasis of NK cells by repressing multiple E-protein target genes including Socs3. Id2 deletion in mature NK cells was incompatible with their homeostasis due to impaired IL-15 receptor signaling and metabolic function and this could be rescued by strong IL-15 receptor stimulation or genetic ablation of Socs3. During NK cell maturation, we observed an inverse correlation between E-protein target genes and Id2. These results shift the current paradigm on the role of ID2, indicating that it is required not only to antagonize E-proteins during NK cell commitment, but constantly required to titrate E-protein activity to regulate NK cell fitness and responsiveness to IL-15.

Lymphoid tissue and plasmacytoid dendritic cells and macrophages do not share a common macrophage-dendritic cell-restricted progenitor.

The relationship between dendritic cells (DCs) and macrophages is often debated. Here we ask whether steady-state, lymphoid-tissue-resident conventional DCs (cDCs), plasmacytoid DCs (pDCs), and macrophages share a common macrophage-DC-restricted precursor (MDP). Using new clonal culture assays combined with adoptive transfer, we found that MDP fractions isolated by previous strategies are dominated by precursors of macrophages and monocytes, include some multipotent precursors of other hematopoietic lineages, but contain few precursors of resident cDCs and pDCs and no detectable common precursors restricted to these DC types and macrophages. Overall we find no evidence for a common restricted MDP leading to both macrophages and FL-dependent, resident cDCs and pDCs.

Safety, infectivity and immunogenicity of a genetically attenuated blood-stage malaria vaccine.

BACKGROUND: There is a clear need for novel approaches to malaria vaccine development. We aimed to develop a genetically attenuated blood-stage vaccine and test its safety, infectivity, and immunogenicity in healthy volunteers. Our approach was to target the gene encoding the knob-associated histidine-rich protein (KAHRP), which is responsible for the assembly of knob structures at the infected erythrocyte surface. Knobs are required for correct display of the polymorphic adhesion ligand P. falciparum erythrocyte membrane protein 1 (PfEMP1), a key virulence determinant encoded by a repertoire of var genes. METHODS: The gene encoding KAHRP was deleted from P. falciparum 3D7 and a master cell bank was produced in accordance with Good Manufacturing Practice. Eight malaria naïve males were intravenously inoculated (day 0) with 1800 (2 subjects), 1.8 × 105 (2 subjects), or 3 × 106 viable parasites (4 subjects). Parasitemia was measured using qPCR; immunogenicity was determined using standard assays. Parasites were rescued into culture for in vitro analyses (genome sequencing, cytoadhesion assays, scanning electron microscopy, var gene expression). RESULTS: None of the subjects who were administered with 1800 or 1.8 × 105 parasites developed parasitemia; 3/4 subjects administered 3× 106 parasites developed significant parasitemia, first detected on days 13, 18, and 22. One of these three subjects developed symptoms of malaria simultaneously with influenza B (day 17; 14,022 parasites/mL); one subject developed mild symptoms on day 28 (19,956 parasites/mL); and one subject remained asymptomatic up to day 35 (5046 parasites/mL). Parasitemia rapidly cleared with artemether/lumefantrine. Parasitemia induced a parasite-specific antibody and cell-mediated immune response. Parasites cultured ex vivo exhibited genotypic and phenotypic properties similar to inoculated parasites, although the var gene expression profile changed during growth in vivo. CONCLUSIONS: This study represents the first clinical investigation of a genetically attenuated blood-stage human malaria vaccine. A P. falciparum 3D7 kahrp- strain was tested in vivo and found to be immunogenic but can lead to patent parasitemia at high doses. TRIAL REGISTRATION: Australian New Zealand Clinical Trials Registry (number: ACTRN12617000824369 ; date: 06 June 2017).

Direct and indirect effects of zinc oxide and titanium dioxide nanoparticles on the decomposition of leaf litter in streams.

As the production of metallic nanoparticles has grown, it is important to assess their impacts on structural and functional components of ecosystems. We investigated the effects of zinc and titanium nanoparticles on leaf decomposition in freshwater habitats. We hypothesized that nanoparticles would inhibit the growth and activity of microbial communities leading to decreased decomposition rates. We also hypothesized that under natural light, the nanoparticles would produce reactive oxygen species that could potentially accelerate decomposition. In the lab, whole Ficus vasta leaves were placed in containers holding one liter of stream water and exposed to either 0, 1, 10 or 100 mg/L of ZnO or TiO2 nanoparticles for six weeks (referred to as Exp. 1). We measured leaf mass loss, microbial metabolism, and bacterial density at 2, 4, and 6 weeks. In a second experiment (referred to as Exp. 2), we measured the effects of light and 10 and 100 mg/L ZnO or TiO2 nanoparticles on leaf mass loss, bacterial density and the bacterial and fungal community diversity over a 2 week period. In Experiment 1, mass loss was significantly reduced at 10 and 100 mg/L after 6 weeks and bacterial density decreased at 100 mg/L. In Experiment 2, there was no effect of ZnO nanoparticles on leaf mass loss, but TiO2 nanoparticles significantly reduced mass loss in the dark but not in the light. One possible explanation is that release of reactive oxygen species by the TiO2 nanoparticles in the light may have increased the rate of leaf decomposition. Bacterial and fungal diversity was highest in the dark, but nanoparticles did not reduce overall diversity.

Living on the edge: biofilms developing in oscillating environmental conditions.

For the first time, the densities and diversity of microorganisms developed on ocean gliders were investigated using flow cytometry and Illumina MiSeq sequencing of 16S and 18S rRNA genes. Ocean gliders are autonomous buoyancy-driven underwater vehicles, equipped with sensors continuously recording physical, chemical, and biological parameters. Microbial biofilms were investigated on unprotected parts of the glider and surfaces coated with base, biocidal and chitosan paints. Biofilms on the glider were exposed to periodical oscillations of salinity, oxygen, temperature, pressure, depth and light, due to periodic ascending and descending of the vehicle. Among the unprotected surfaces, the highest microbial abundance was observed on the bottom of the glider's body, while the lowest density was recorded on the glider's nose. Antifouling paints had the lowest densities of microorganisms. Multidimensional analysis showed that the microbial communities formed on unprotected parts of the glider were significantly different from those on biocidal paint and in seawater.

Identification of Novel Human NK Cell Progenitor Subsets.

Understanding the pathways and regulation of human haematopoiesis, in particular, lymphopoiesis, is vital to manipulation of these processes for therapeutic purposes. However, although haematopoiesis has been extensively characterised in mice, translation of these findings to human biology remains rudimentary. Here, we describe the isolation of three progenitor subsets from human foetal bone marrow that represent differential stages of commitment to the natural killer (NK) cell lineage based on IL-15 responsiveness. We identify CD7 as a marker of IL-15 responsive progenitors in human bone marrow and find that this expression is maintained throughout commitment and maturation. Within the CD7⁺ fraction, we focussed on the lineage potential of three subsets based on CD127 and CD117 expression and observed restricted lymphoid and biased NK cell potential amongst subsets. We further demonstrate the presence of subsets similar in both phenotype and function in umbilical cord blood and the bone marrow of humanised mice, validating these as appropriate sources of progenitors for the investigation of human haematopoiesis. Overall, we describe several stages in the process of lymphopoiesis that will form the basis of investigating the regulators of this process in humans.

Self-decontaminating photocatalytic zinc oxide nanorod coatings for prevention of marine microfouling: a mesocosm study.

The antifouling (AF) properties of zinc oxide (ZnO) nanorod coated glass substrata were investigated in an out-door mesocosm experiment under natural sunlight (14:10 light: dark photoperiod) over a period of five days. The total bacterial density (a six-fold reduction) and viability (a three-fold reduction) was significantly reduced by nanocoatings in the presence of sunlight. In the absence of sunlight, coated and control substrata were colonized equally by bacteria. MiSeq Illumina sequencing of 16S rRNA genes revealed distinct bacterial communities on the nanocoated and control substrata in the presence and absence of light. Diatom communities also varied on nanocoated substrata in the presence and the absence of light. The observed AF activity of the ZnO nanocoatings is attributed to the formation of reactive oxygen species (ROS) through photocatalysis in the presence of sunlight. These nanocoatings are a significant step towards the production of an environmentally friendly AF coating that utilizes a sustainable supply of sunlight.

Convergent differentiation: myeloid and lymphoid pathways to murine plasmacytoid dendritic cells.

The developmental origin of IFN-producing plasmacytoid dendritic cells (pDCs) has been uncertain. In the present study, we tracked the development of pDCs in cultures of BM precursors stimulated with Flt3 ligand. Common myeloid precursors (CMPs) produced both conventional DCs (cDCs) and pDCs via the DC-restricted common DC precursor. Common lymphoid precursors (CLPs) produced only a few cDCs with variable efficiency, but produced pDCs via a transient intermediate precursor with B-cell potential. The pDCs of both origins produced IFN-α when stimulated with CpG oligonucleotides. The pDCs of CLP origin showed evidence of past RAG1 expression and had D-J rearrangements in IgH genes. Most pDCs and all cDCs of CMP origin lacked these signs of a lymphoid past. However, in these cultures, some pDCs of CMP origin showed evidence of past RAG1 expression and had D-J IgH gene rearrangements; most of these derived from a subset of CMPs already expressing RAG1.

The acquisition of antigen cross-presentation function by newly formed dendritic cells.

The development of Ag-presenting functions by murine dendritic cells (DCs) of the CD8(+) DC lineage was studied using a Flt-3 ligand stimulated bone-marrow culture system. Although newly formed DCs of this lineage are capable of Ag uptake and efficient presentation to T cells on MHC class II, they initially lack the ability to cross-present exogenous Ags on MHC class I. Cross-presentation capacity is acquired as a subsequent maturation step, promoted by cytokines such as GM-CSF. The development of cross-presentation capacity by the DCs in these cultures may be monitored by the parallel development of DC surface expression of CD103. However, the expression of CD103 and cross-presentation capacity are not always linked; therefore, CD103 is not an essential part of the cross-presentation machinery. These results explain the considerable variability in CD103 expression by CD8(+) DCs as well as the findings that not all DCs of this lineage are capable of cross-presentation.

Comparison of intraocular lens power prediction accuracy of formulas in American Society of Cataract and Refractive Surgery post-refractive surgery calculator in eyes with prior radial keratotomy.

Purpose: To evaluate the accuracy of intraocular lens (IOL) power prediction of the formulas available on the American Society of Cataract and Refractive Surgery (ASCRS) post-refractive calculator in eyes with prior radial keratotomy (RK) for myopia. Methods: This retrospective study included 25 eyes of 18 patients whose status was post-RK for treatment of myopia, which had undergone cataract extraction with IOL implantation. Prediction error was calculated as the difference between implanted IOL power and predicted power by various formulae available on ASCRS post-refractive calculator. The formulas compared were Humphrey Atlas method, IOLMaster/Lenstar method, Barrett True-K no-history formula, ASCRS Average power, and ASCRS Maximum power on ASCRS post-refractive calculator. Results: Median absolute errors were the least for Barrett True-K and ASCRS Maximum power, that is, 0.56 (0.25, 1.04) and 0.56 (0.25, 1.06) D, respectively, and that of Atlas method was 1.60 (0.85, 2.28) D. Median arithmetic errors were positive for Atlas, Barrett True-K, ASCRS Average (0.86 [-0.17, 1.61], 0.14 [-0.22 to 0.54], and 0.23 [-0.054, 0.76] D, respectively) and negative for IOLMaster/Lenstar method and ASCRS Maximum power (-0.02 [-0.46 to 0.38] and - 0.48 [-1.06 to - 0.22] D, respectively). Multiple comparison analysis of Friedman's test revealed that Atlas formula was significantly different from IOLMaster/Lenstar, Barrett True-K, and ASCRS Maximum power; ASCRS Maximum power was significantly different from all others (P < 0.00001). Conclusion: In post-RK eyes, Barrett True-K no-history formula and ASCRS Maximum power given by the ASCRS calculator were more accurate than other available formulas, with ASCRS Maximum leading to more myopic outcomes when compared to others.

Impact of crossplay between ocular aberrations and depth of focus in topo-guided laser-assisted in situ keratomileusis outcomes.

Purpose: To develop a nomogram in cases with mismatch between subjective and Topolyzer cylinder, and based on the magnitude of the mismatch, customize a treatment plan to attain good visual outcomes post-laser-assisted in situ keratomileusis (LASIK) surgery. Methods: The patients were evaluated preoperatively using corneal tomography with Pentacam. Five optimal corneal topography scans were obtained from the Topolyzer Vario were used for planning the LASIK treatment. For the nomogram purpose, the patients were divided into three categories based on the difference between the subjective cylinder and Topolyzer (corneal) cylinder. The first group (group 1) consisted of eyes of patients, where the difference was less than or equal to 0.4 D. The second group (group 2) consisted of eyes, where the difference was more than 0.4 D and the subjective cylinder was lesser than the Topolyzer cylinder. The third group (group 3) included eyes where the difference was more than 0.4 D but the subjective cylinder was greater than the Topolyzer cylinder. LASIK was performed with the WaveLight FS 200 femtosecond laser and WaveLight EX500 excimer laser. Assessment of astigmatism correction for the three groups was done using Aplins vector analysis. For comparison of proportions, Chi-square test was used. A P value less than 0.05 was considered statistically significant. Results: The UDVA was statistically significantly different when compared between groups 1 and 2 (P = 0.02). However, the corrected distance visual acuity (CDVA) was similar among all the three groups (P = 0.1). Group 3 showed an increase of residual cylinder by -0.25 D, which was significant at intermediate and near reading distances (P < 0.05). Group 3 showed significantly higher target-induced astigmatism (TIA) compared to groups 1 and 2 (P = 0.01). The mean surgically induced astigmatism (SIA) was the least in group 2, which was statistically significant (P < 0.01). Conclusion: The outcomes for distance vision using our nomogram postoperatively were excellent, but further refinement for improving the near vision outcomes is required.

The C-type lectin Clec12A present on mouse and human dendritic cells can serve as a target for antigen delivery and enhancement of antibody responses.

We have cloned the mouse and human C-type lectin Clec12A, expressed both, and produced mAb recognizing both. Mouse Clec12A is highly expressed on splenic CD8(+) dendritic cells (DC) and plasmacytoid DC. A proportion of CD8(-)DC also expresses lower levels of Clec12A, as do monocytes, macrophages, and B cells. Human CLEC12A, like the mouse counterpart, is expressed on blood monocytes and DC, including pDC and BDCA-3(+)DC, the proposed equivalent of mouse CD8(+)DC. To determine whether Ag targeted to Clec12A could induce immune responses, mice were injected with a rat mAb recognizing Clec12A, or a control rat mAb, then production of anti-rat Ig was measured. Anti-Clec12A mAb alone produced only moderate responses, but these were amplified by coinjecting only small amounts of LPS as a DC activation agent. Furthermore, when OVA was conjugated to anti-Clec12A mAb, OVA-specific T cells were induced to proliferate. This Ag presentation to naive T cells was due to targeting conventional DC, because their ablation eliminated T cell activation. The potent Ab responses induced using microgram amounts of anti-Clec12A and minimal amounts of adjuvant demonstrate that this molecule can be used as an Ag-delivery target to enhance Ab responses to vaccines.

Disinfection of Bacteria in Water by Capacitive Deionization.

Clean water is one of the primary UN sustainable development goals for 2,030 and sustainable water deionization and disinfection is the backbone of that goal. Capacitive deionization (CDI) is an upcoming technique for water deionization and has shown substantial promise for large scale commercialization. In this study, activated carbon cloth (ACC) electrode based CDI devices are used to study the removal of ionic contaminants in water and the effect of ion concentrations on the electrosorption and disinfection functions of the CDI device for mixed microbial communities in groundwater and a model bacterial strain Escherichia coli. Up to 75 % of microbial cells could be removed in a single pass through the CDI unit for both synthetic and groundwater, while maintaining the salt removal activity. Mortality of the microbial cells were also observed during the CDI cell regeneration and correlated with the chloride ion concentrations. The power consumption and salt removal capacity in the presence and absence of salt were mapped and shown to be as low as 0.1 kWh m-3 and 9.5 mg g-1, respectively. The results indicate that CDI could be a viable option for single step deionization and microbial disinfection of brackish water.

Toxicity of Different Zinc Oxide Nanomaterials at 3 Trophic Levels: Implications for Development of Low-Toxicity Antifouling Agents.

Because zinc oxide (ZnO) nanomaterials are used in antifouling and antibacterial solutions, understanding their toxic effects on different aquatic organisms is essential. In the present study, we evaluated the toxicity of ZnO nanoparticles of 10 to 30 nm (ZnONPI) and 80 to 200 nm (ZnONPII), ZnO nanorods (width 80 nm, height 1.7 µm) attached to the support substrate (glass, ZnONRG) and not attached (ZnONRS), as well as Zn2+ ions at concentrations ranging from 0.5 to 100 mg/L. Toxicity was evaluated using the microalga Dunaliella salina, the brine shrimp Artemia salina, and the marine bacterium Bacillus cereus. The highest toxicity was observed for ZnONPs (median lethal concentration [LC50] ~15 mg/L) and Zn2+ ions (LC50 ~13 mg/L), whereas the lowest toxicity found for ZnO nanorods (ZnONRG LC50 ~60 mg/L; ZnONRS LC50 ~42 mg/L). The presence of the support substrate in case of ZnO nanorods reduced the associated toxicity to aquatic organisms. Smaller ZnONPs resulted in the highest Zn2+ ion dissolution among tested nanostructures. Different aquatic organisms responded differently to ZnO nanomaterials, with D. salina and B. cereus being more sensitive than A. salina. Toxicity of nanostructures increased with an increase of the dose and the time of exposure. Supported ZnO nanorods can be used as a low-toxicity alternative for future antimicrobial and antifouling applications. Environ Toxicol Chem 2020;39:1343-1354. © 2020 SETAC.

Antimicrobial Activity Enhancement of Poly(ether sulfone) Membranes by in Situ Growth of ZnO Nanorods.

Composite poly(ether sulfone) membranes integrated with ZnO nanostructures either directly blended or grown in situ have enhanced antibacterial activity with improved functionality in reducing the biofouling in water treatment applications. The pore structure and surface properties of the composite were studied to investigate the effect of the addition of ZnO nanostructures. The hydrophilicity of the blended membranes increased with a higher content of ZnO nanoparticles in the membrane (2-6%), which could be further controlled by varying the growth conditions of ZnO nanorods on the polymer surface. Improved water flux, bovine serum albumin rejection, and inhibition of Escherichia coli bacterial growth under visible light irradiation was observed for the membranes decorated with ZnO nanorods compared to those in the membranes simply blended with ZnO nanoparticles. No regrowth of E. coli was recorded even 2 days after the incubation.

Bioinspired nanocoatings for biofouling prevention by photocatalytic redox reactions.

Aquaculture is a billion dollar industry and biofouling of aquaculture installations has heavy economic penalties. The natural antifouling (AF) defence mechanism of some seaweed that inhibits biofouling by production of reactive oxygen species (ROS) inspired us to mimic this process by fabricating ZnO photocatalytic nanocoating. AF activity of fishing nets modified with ZnO nanocoating was compared with uncoated nets (control) and nets painted with copper-based AF paint. One month experiment in tropical waters showed that nanocoatings reduce abundances of microfouling organisms by 3-fold compared to the control and had higher antifouling performance over AF paint. Metagenomic analysis of prokaryotic and eukaryotic fouling organisms using next generation sequencing platform proved that nanocoatings compared to AF paint were not selectively enriching communities with the resistant and pathogenic species. The proposed bio-inspired nanocoating is an important contribution towards environmentally friendly AF technologies for aquaculture.

Innate immunodeficiency following genetic ablation of Mcl1 in natural killer cells.

The cytokine IL-15 is required for natural killer (NK) cell homeostasis; however, the intrinsic mechanism governing this requirement remains unexplored. Here we identify the absolute requirement for myeloid cell leukaemia sequence-1 (Mcl1) in the sustained survival of NK cells in vivo. Mcl1 is highly expressed in NK cells and regulated by IL-15 in a dose-dependent manner via STAT5 phosphorylation and subsequent binding to the 3'-UTR of Mcl1. Specific deletion of Mcl1 in NK cells results in the absolute loss of NK cells from all tissues owing to a failure to antagonize pro-apoptotic proteins in the outer mitochondrial membrane. This NK lymphopenia results in mice succumbing to multiorgan melanoma metastases, being permissive to allogeneic transplantation and being resistant to toxic shock following polymicrobial sepsis challenge. These results clearly demonstrate a non-redundant pathway linking IL-15 to Mcl1 in the maintenance of NK cells and innate immune responses in vivo.

Plasmacytoid dendritic cell development.

Interferon-producing plasmacytoid dendritic cells (pDC) are a specialized branch of the dendritic cell (DC) family, and their differentiation in mice is closely linked to that of conventional DC (cDC). Several different developmental pathways retain the potential to form pDC and are likely to contribute to the steady-state pDC population. A lymphoid pathway to DC development produces mainly pDC as a branch otherwise leading to B-cell development; such pDC may carry relics of a lymphoid past such as DJ rearrangements of immunoglobulin heavy chain (IgH) genes. The myeloid pathway to pDC and cDC is better known, but recent reassessment has revealed several substreams of development with separate DC-committed precursors. One substream has a lymphoid-like aspect, involving a precursor expressing RAG-1 and producing pDC with IgH gene rearrangements. Another more biased to cDC production produces pDC without such IgH gene rearrangements. Finally, there is the production of interferon-producing pDC-like cells that are not pDC but appear to be cDC precursors; these do not express key pDC markers such as CCR9. Initiation of the DC and then the pDC developmental program overrides any surface marker-expressed developmental bias to other myeloid or lymphoid lineages, resulting in an apparent convergent differentiation to the pDC form. A DC fate is sometimes imprinted early in development, upstream of identifiable myeloid, or lymphoid precursors. This suggests that DC, including pDC, represent a distinct hematopoietic lineage separate from conventional myeloid or lymphoid cells.