Tree Age Distributions Reveal Large-Scale Disturbance-Recovery Cycles in Three Tropical Forests

Over the past few decades there has been a growing realization that a large share of apparently ‘virgin’ or ‘old-growth’ tropical forests carries a legacy of past natural or anthropogenic disturbances that have a substantial effect on present-day forest composition, structure and dynamics. Yet, direct evidence of such disturbances is scarce and comparisons of disturbance dynamics across regions even more so. Here we present a tree-ring based reconstruction of disturbance histories from three tropical forest sites in Bolivia, Cameroon, and Thailand. We studied temporal patterns in tree regeneration of shade-intolerant tree species, because establishment of these trees is indicative for canopy disturbance. In three large areas (140–300 ha), stem disks and increment cores were collected for a total of 1154 trees (>5 cm diameter) from 12 tree species to estimate the age of every tree. Using these age estimates we produced population age distributions, which were analyzed for evidence of past disturbance. Our approach allowed us to reconstruct patterns of tree establishment over a period of around 250 years. In Bolivia, we found continuous regeneration rates of three species and a peaked age distribution of a long-lived pioneer species. In both Cameroon and Thailand we found irregular age distributions, indicating strongly reduced regeneration rates over a period of 10–60 years. Past fires, windthrow events or anthropogenic disturbances all provide plausible explanations for the reported variation in tree age across the three sites. Our results support the recent idea that the long-term dynamics of tropical forests are impacted by large-scale disturbance-recovery cycles, similar to those driving temperate forest dynamics.This study was financially supported by the European Research Council (ERC, grant #242955). The paper is derived from the work done in partial fulfillment of the requirements for a PhD degree of the first author (MV) at Wageningen University (Vlam, 2014)S

Potential sources of bias in the climate sensitivities of fish otolith biochronologies

Analysis of growth increments in the hard parts of animals (e.g., fish otoliths) can be used to assess how organisms respond to variability in environmental conditions. In this study, mixed-effects models were applied to otolith data simulated for two hypothetical fish populations with assumed biological parameters and known growth response to environmental variability. Our objective was to assess the sensitivity of environment–growth relationships derived from otolith biochronologies when challenged with a range of realistic ageing errors and sampling regimes. We found that the development of a robust biochronology and the precision of environmental effect estimates can be seriously hampered by insufficient sample size. Moreover, the introduction of even moderate ageing error into the data can cause substantial underestimation of environmental sources of growth variation. This underestimation diminished our capacity to correctly quantify the known environment–growth relationship and more generally will lead to overly conservative conclusions concerning the growth response to environmental change. Careful study design, reduction of ageing errors, and large sample sizes are critical prerequisites if robust inferences are to be made from biochronological data.publishedVersio

Aberrant phenotype of circulating antigen presenting cells in giant cell arteritis and polymyalgia rheumatica

BACKGROUND: Giant Cell Arteritis (GCA) and Polymyalgia Rheumatica (PMR) are overlapping inflammatory diseases. Antigen-presenting cells (APCs), including monocytes and dendritic cells (DCs), are main contributors to the immunopathology of GCA and PMR. However, little is known about APC phenotypes in the peripheral blood at the time of GCA/PMR diagnosis.METHODS: APCs among peripheral blood mononuclear cells (PBMCs) of treatment-naive GCA and PMR patients were compared to those in age- and sex-matched healthy controls (HCs) using flow cytometry (n=15 in each group). We identified three monocyte subsets, and three DC subsets: plasmacytoid DCs (pDCs), CD141+ conventional DCs (cDC1) and CD1c+ conventional DCs (cDC2). Each of these subsets was analyzed for expression of pattern recognition receptors (TLR2, TLR4), immune checkpoints (CD86, PDL1, CD40) and activation markers (HLA-DR, CD11c).RESULTS: t-SNE plots revealed a differential clustering of APCs between GCA/PMR and HCs. Further analyses showed shifts in monocyte subsets and a lower proportion of the small population of cDC1 cells in GCA/PMR, whereas cDC2 proportions correlated negatively with CRP (r=-0.52). Classical monocytes of GCA/PMR patients show reduced expression of TLR2, HLA-DR, CD11c, which was in contrast to non-classical monocytes that showed higher marker expression. Additionally, single cell RNA sequencing in GCA patients identified a number of differentially expressed genes related to inflammation and metabolism in APCs.CONCLUSION: Circulating non-classical monocytes display an activated phenotype in GCA/PMR patients at diagnosis, whereas classical monocytes show reduced expression of activation markers. Whether these findings reflect APC migration patterns or the effects of long-term inflammation remains to be investigated.</p

Functionally Heterogenous Macrophage Subsets in the Pathogenesis of Giant Cell Arteritis:Novel Targets for Disease Monitoring and Treatment

Giant cell arteritis (GCA) is a granulomatous large-vessel vasculitis that affects adults above 50 years of age. In GCA, circulating monocytes are recruited to the inflamed arteries. With cues from the vascular microenvironment, they differentiate into macrophages and play important roles in the pathogenesis of GCA via pro-inflammatory cytokine production and vascular remodeling. However, a deeper understanding of macrophage heterogeneity in GCA pathogenesis is needed to assist the development of novel diagnostic tools and targeted therapies. Here, we review the current knowledge on macrophage heterogeneity and diverse functions of macrophage subsets in the pathogenesis of GCA. We next discuss the possibility to exploit their heterogeneity as a source of novel biomarkers and as targets for nuclear imaging. Finally, we discuss novel macrophage-targeted therapies and future directions for targeting these cells in GCA

A Distinct Macrophage Subset Mediating Tissue Destruction and Neovascularization in Giant Cell Arteritis:Implication of the YKL-40/Interleukin-13 Receptor alpha 2 Axis

OBJECTIVE: Macrophages mediate inflammation, angiogenesis and tissue destruction in giant cell arteritis (GCA). Serum levels of the macrophage-associated protein YKL-40 (chitinase-3 like-1), previously linked to angiogenesis and tissue remodeling, remain elevated in GCA despite glucocorticoid treatment. Here, we aimed to investigate the contribution of YKL-40 to vasculopathy in GCA. METHODS: Immunohistochemistry was performed on GCA temporal artery biopsies (TABs; n=12) and aortas (n=10) for detection of YKL-40, its receptor IL-13Rα2, macrophage markers PU.1 and CD206, and the tissue-destructive protein MMP-9. Ten non-inflamed TABs served as controls. In vitro experiments with GM-CSF- or M-CSF-skewed monocyte-derived macrophages (GM-MØs or M-MØs) were conducted to study the dynamics of YKL-40 production. Next, silencing RNA (siRNA)-mediated knock-down of YKL-40 in GM-MØs was performed to study its effect on MMP-9 production. Finally, the angiogenic potential of YKL-40 was investigated by tube formation experiments using human microvascular endothelial cells (HMVECs). RESULTS: YKL-40 was abundantly expressed by a CD206+MMP-9+ macrophage subset in inflamed TABs and aortas. GM-MØs of GCA patients, but not of healthy controls, released significantly higher levels of YKL-40, compared to M-MØs. In inflamed TABs, IL-13Rα2 was expressed by macrophages and endothelial cells. Functionally, knock-down of YKL-40 led to a 10-50% reduction in MMP-9 production by macrophages, whereas exposure of HMVECS to YKL-40 led to significantly increased tube formation. CONCLUSION: In GCA, a GM-CSF-skewed, CD206+MMP9+ macrophage subset expresses high levels of YKL-40 which may stimulate tissue destruction and angiogenesis through IL-13Rα2 signaling. Targeting YKL-40 or GM-CSF may inhibit macrophages that are currently insufficiently suppressed by glucocorticoids

Distinct macrophage phenotypes skewed by local granulocyte macrophage colony-stimulating factor (GM-CSF) and macrophage colony-stimulating factor (M-CSF) are associated with tissue destruction and intimal hyperplasia in giant cell arteritis

Objective: To determine the presence and spatial distribution of different macrophage phenotypes, governed by granulocyte macrophage colony-stimulating factor (GM-CSF) and macrophage colony-stimulating factor (M-CSF) skewing signals, in giant cell arteritis (GCA) lesions. Methods: Temporal artery biopsies (TABs, n = 11) from treatment-naive GCA patients, aorta samples from GCA-related aneurysms (n = 10) and atherosclerosis (n = 10) were stained by immunohistochemistry targeting selected macrophage phenotypic markers, cytokines, matrix metalloproteinases (MMPs) and growth factors. In vitro macrophage differentiation (n = 10) followed by flow cytometry, Luminex assay and ELISA were performed to assess whether GM-CSF and M-CSF are drivers of macrophage phenotypic heterogeneity. Results: A distinct spatial distribution pattern of macrophage phenotypes in TABs was identified. CD206+/MMP-9+ macrophages were located at the site of tissue destruction, whereas FRβ+ macrophages were located in the inner intima of arteries with high degrees of intimal hyperplasia. Notably, this pattern was also observed in macrophage-rich areas in GCA aortas but not in atherosclerotic aortas. Flow cytometry showed that GM-CSF treatment highly upregulated CD206 expression, while FRβ was expressed by M-CSF-skewed macrophages, only. Furthermore, localised expression of GM-CSF and M-CSF was detected, likely contributing to macrophage heterogeneity in the vascular wall. Conclusions: Our data document a distinct spatial distribution pattern of CD206+/MMP-9+ macrophages and FRβ+ macrophages in GCA linked to tissue destruction and intimal proliferation, respectively. We suggest that these distinct macrophage phenotypes are skewed by sequential GM-CSF and M-CSF signals. Our study adds to a better understanding of the development and functional role of macrophage phenotypes in the pathogenesis of GCA and opens opportunities for the design of macrophage-targeted therapies

Decreased Expression of Negative Immune Checkpoint VISTA by CD4+T Cells Facilitates T Helper 1, T Helper 17, and T Follicular Helper Lineage Differentiation in GCA

Loss of immune checkpoint (IC) Programmed Death-1 (PD-1) and PD-Ligand1 (PD-L1) expression has been implicated in the immunopathology of Giant Cell Arteritis (GCA). The contribution of the negative immune checkpoint V-domain Immunoglobulin-containing suppressor of T cell activation (VISTA) to GCA pathology has not yet been studied. The aim of our study was to investigate if expression of VISTA and other IC molecules by peripheral blood (PB) immune cells is modulated in GCA and at the site of vascular inflammation. In addition, we assessed the effect of VISTA-Ig engagement on in vitro CD4+ T helper (Th) lineage differentiation. To this end, frequencies of monocytes expressing CD80/86, PD-L1, PD-L2, and VISTA were determined in blood samples from 30 GCA patients and 18 matched healthy controls by flow cytometry. In parallel, frequencies of CD4+ cells expressing CD28, Cytotoxic T-Lymphocyte-associated antigen-4 (CTLA-4), PD-1, and VISTA were determined. Immunohistochemistry was employed to detect VISTA, PD-1, and PD-L1-expressing cells in temporal artery biopsies (TABs) diagnostic of GCA. Furthermore, the effect of VISTA-Ig on in vitro CD4+ Th lineage differentiation in patients and controls was determined. Our study shows that frequencies of CD80/CD86+ and VISTA+ monocytes were decreased in treated GCA patients only. Moreover, proportions of PD-1+ and VISTA+ Th cells were significantly decreased in GCA patients. Clear infiltration of VISTA+, PD1+, and PD-L1+ cells was seen in GCA TABs. Finally, VISTA-Ig engagement failed to suppress Th1, Th17, and Tfh lineage development in GCA. Our results indicate that decreased expression of VISTA may facilitate development of pathogenic Th1 and Th17 cells in GCA