Thrombopoietin cooperates with FLT3-ligand in the generation of plasmacytoid dendritic cell precursors from human hematopoietic progenitors

Type 1 interferon-producing cells (IPCs), also known as plasmacytoid dendritic cell (DC) precursors, represent the key effectors in antiviral innate immunity and triggers for adaptive immune responses. IPCs play important roles in the pathogenesis of systemic lupus erythematosus (SLE) and in modulating immune responses after hematopoietic stem cell transplantation. Understanding IPC development from hematopoietic progenitor cells (HPCs) may provide critical information in controlling viral infection, autoimmune SLE, and graft-versus-host disease. FLT3-ligand (FLT3-L) represents a key IPC differentiation factor from HPCs. Although hematopoietic cytokines such as interleukin-3 (IL-3), IL-7, stem cell factor (SCF), macrophage-colony-stimulating factor (M-CSF), and granulocyte M-CSF (GM-CSF) promote the expansion of CD34+ HPCs in FLT3-L culture, they strongly inhibit HPC differentiation into IPCs. Here we show that thrombopoietin (TPO) cooperates with FLT3-L, inducing CD34+ HPCs to undergo a 400-fold expansion in cell numbers and to generate more than 6 x 10(6) IPCs per 10(6) CD34+ HPCs within 30 days in culture. IPCs derived from HPCs in FLT3-L/TPO cultures display blood IPC phenotype and have the capacity to produce large amounts of interferon-alpha (IFN-alpha) and to differentiate into mature DCs. This culture system, combined with the use of adult peripheral blood CD34+ HPCs purified from G-CSF-mobilized donors, permits the generation of more than 10(9) IPCs from a single blood donor.     

Ancient plant DNA reveals High Arctic greening during the Last Interglacial

Summer warming is driving a greening trend across the Arctic, with the potential for large-scale amplification of climate change due to vegetation-related feedbacks [Pearson et al., Nat. Clim. Chang. (3), 673–677 (2013)]. Because observational records are sparse and temporally limited, past episodes of Arctic warming can help elucidate the magnitude of vegetation response to temperature change. The Last Interglacial ([LIG], 129,000 to 116,000 y ago) was the most recent episode of Arctic warming on par with predicted 21st century temperature change [Otto-Bliesner et al., Philos. Trans. A Math. Phys. Eng. Sci. (371), 20130097 (2013) and Post et al., Sci. Adv. (5), eaaw9883 (2019)]. However, high-latitude terrestrial records from this period are rare, so LIG vegetation distributions are incompletely known. Pollen-based vegetation reconstructions can be biased by long-distance pollen transport, further obscuring the paleoenvironmental record. Here, we present a LIG vegetation record based on ancient DNA in lake sediment and compare it with fossil pollen. Comprehensive plant community reconstructions through the last and current interglacial (the Holocene) on Baffin Island, Arctic Canada, reveal coherent climate-driven community shifts across both interglacials. Peak LIG warmth featured a ∼400-km northward range shift of dwarf birch, a key woody shrub that is again expanding northward. Greening of the High Arctic—documented here by multiple proxies—likely represented a strong positive feedback on high-latitude LIG warming. Authenticated ancient DNA from this lake sediment also extends the useful preservation window for the technique and highlights the utility of combining traditional and molecular approaches for gleaning paleoenvironmental insights to better anticipate a warmer future.

The Arctic is greening as shrub biomass increases and vegetation ranges shift north in response to summer warming (1, 2). This process—one of the clearest terrestrial manifestations of climate change thus far—has major implications both for local ecosystems and for global energy balance and biogeochemical systems (3–5). In particular, taller shrubs darken otherwise snow-covered surfaces, contributing to the albedo feedback (6, 7), and enhanced evapotranspiration is expected to result in a positive greenhouse feedback (8). Shrub cover also impacts soil thermal regime, which may impact permafrost vulnerability (9–11). Because feedbacks related to Arctic greening are complex and potentially large in magnitude, estimating the extent and rate of northward shrub migration is a vital component of predicting future warming.Past warm periods serve as valuable analogs for understanding the extent of Arctic greening under well-constrained climate conditions. The Last Interglacial (LIG; Marine Isotope Stage [MIS] 5e, 129 to 116 ka [thousands of years before present]) was ∼1 °C warmer than the preindustrial period globally, but the Arctic experienced amplified warming due to higher summer insolation anomalies and positive feedbacks at high latitudes (12, 13). The Eastern Canadian Arctic and Greenland, in particular, were likely ∼4 to 8 °C warmer in summer than present (Fig. 1) (14–18). LIG sediment records from this region thus provide an archive of the vegetation response to Arctic warming at levels comparable to predicted 21^st-century climate change (19).Open in a separate windowFig. 1.Map of Baffin Island and Lake CF8 study area. The symbols represent maximum LIG temperature anomalies based on terrestrial proxy records (shape indicates proxy type) from Baffin Island and Greenland (see SI Appendix, Table S1 for metadata). The shaded regions indicate Arctic bioclimate subzones delineations (29), including modern Betula range in subzones D and E. We note that a small outlier population of Betula occurs east of the D/E boundary on Baffin Island (not captured by vegetation map resolution) (38).While most High Arctic lake basins were scraped clean by ice sheet erosion during the last glaciation and thus only contain postglacial sediments, lakes with small, low-relief catchments within regions of cold-based, slow-flowing ice were protected from erosion. Several such sites have been discovered on eastern Baffin Island, Arctic Canada and contain stratified records of multiple interglacials (20–22). Previous work from Lake CF8 on northeastern Baffin Island (Fig. 1 and SI Appendix, Fig. S1) demonstrates that its sediment record spans at least three interglacials (∼200 ka), including a substantially warmer-than-present LIG as indicated by chironomids, diatoms, and geochemical proxies (15, 23).We targeted the multi-interglacial record from Lake CF8 to assess the vegetation response to pronounced warmth during the LIG and moderate warmth during the Holocene. Pollen produced by some key shrubs and trees, including Betula (birch), is efficiently wind-transported and thus present in lake sediments far north of their ranges (24, 25). We therefore analyzed both sedimentary ancient DNA (sedaDNA), which is sourced locally from within the lake catchment and does not include pollen-derived DNA (26), and fossil pollen to generate a robust vegetation record spanning the last ∼130 ka. Taken together, DNA-inferred plant communities and pollen-inferred July air temperatures provide insight into Arctic plant range shifts under strong summer warming.     

Bilirubin protects astrocytes from its own toxicity by inducing up-regulation and translocation of multidrug resistance-associated protein 1 (Mrp1)

Unconjugated bilirubin (UCB) causes encephalopathy in severely jaundiced neonates by damaging astrocytes and neurons. Astrocytes, which help defend the brain against cytotoxic insults, express the ATP-dependent transporter, multidrug resistance-associated protein 1 (Mrp1), which mediates export of organic anions, probably including UCB. We therefore studied whether exposure to UCB affects the expression and intracellular localization of Mrp1 in cultured mouse astroglial cells (>95% astrocytes). Mrp1 was localized and quantitated by confocal laser scanning microscopy and double immunofluorescence labeling by using specific antibodies against Mrp1 and the astrocyte marker glial fibrillary acidic protein, plus the Golgi marker wheat germ agglutinin (WGA). In unexposed astrocytes, Mrp1 colocalized with WGA in the Golgi apparatus. Exposure to UCB at a low unbound concentration (Bf) of 40 nM caused rapid redistribution of Mrp1 from the Golgi throughout the cytoplasm to the plasma membrane, with a peak 5-fold increase in Mrp1 immunofluorescence intensity from 30 to 120 min. Bf above aqueous saturation produced a similar but aborted response. Exposure to this higher Bf for 16 h markedly decreased Trypan blue exclusion and methylthiazoletetrazoilum activity and increased apoptosis 5-fold by terminal deoxynucleotidyltransferase-mediated dUTP nick end labeling assay. These toxic effects were modestly increased by inhibition of Mrp1 activity with 3-([3-(2-[7-chloro-2-quinolinyl]ethenyl)phenyl-(3-dimethylamino-3-oxopropyl)-thio-methyl]thio)propanoic acid (MK571). By contrast, Bf=40 nM caused injury only if Mrp1 activity was inhibited by MK571, which also blocked translocation of Mrp1. Our conclusion is that in astrocytes, UCB up-regulates expression of Mrp1 and promotes its trafficking from the Golgi to the plasma membrane, thus moderating cytotoxicity from UCB, presumably by limiting its intracellular accumulation.     

Intra- and intermuscular variations of postmortem protein degradation for PMI estimation

International Journal of Legal Medicine - In recent years, protein decomposition has become of increasing interest for the use in forensic estimation of the postmortem interval (PMI). Especially...     

Urokinase-type plasminogen activator (uPA) decreases hepatic SR-BI expression and impairs HDL-mediated reverse cholesterol transport

Objectives

The aim of the present study was to investigate the effect of urokinase-type plasminogen activator (uPA) on the expression of the scavenger receptor class B type I (SR-BI) in hepatocytes, and its impact on the removal of HDL-cholesteryl ester (CE) in the liver.

Methods and results

Huh7 hepatoma cell lines were incubated with increasing concentrations of uPA. uPA dose-dependently decreased SR-BI protein expression, as determined by flow cytometry (FACS) and by Western blot assays, and down-regulated SR-BI gene expression. Functionally, uPA decreased both the cellular binding of HDL to Huh7 hepatocytes, and the selective uptake of CE from HDL, as determined by several methods including BODIPY staining, cellular cholesterol determination and chasing radio-labeled CE transfer from HDL to the cells. These results were further confirmed using primary rat hepatocyes. The effect of uPA on hepatic SR-BI expression was mediated via binding to the uPA receptor (uPAR). In vivo, SR-BI protein and gene expressions were found to be increased in hepatocytes derived from the uPAR-KO mice compared to C57Bl/6 mice, and in parallel HDL-cholesterol levels in plasma derived from uPAR-KO mice were decreased. Moreover, deficiency of uPAR significantly accelerated the plasma decay of injected HDL-[³H]CE.

Conclusions

The results of this study suggest that uPA decreases the removal of HDL-CE in the liver via suppression of the hepatic SR-BI expression. Impaired reverse cholesterol transport (RCT) may result in atherogenic dysfunctional HDL metabolism and may contribute to atherosclerosis development.