Functional Niche Research Articles

Response of chestnut hybrid seedlings to forest management strategies in an Appalachian cove forest ecosystem

American chestnut (Castanea dentata (Marsh) Borkh.), once a co-dominant canopy species in eastern U.S. forests, has been functionally extinct for almost a century. Hybrid American-Chinese chestnuts, with the potential to be blight-resistant, present an opportunity to study its functional niche. The objective of this research was to identify silvicultural strategies related to light, herbivory, and competition for increasing the success of hybrid chestnut (BC3F3 generation) plantings in an Appalachian cove forest ecosystem in West Virginia (WV). Large and small circular canopy gaps were created to manipulate light with the prediction that seedlings would perform best in larger gaps. Within these gaps, landscape fabric was added to half of the seedlings to decrease vegetative competition and 30.5 cm tall tree shelters were placed around half of the seedlings to help mitigate rodent predation. After five years, results show an interaction between landscape fabric and gap size whereby landscape fabric had a positive effect on seedling height in large gaps but not in small gaps (p < 0.05). Survival was dependent on interactions between treatments with the highest seedling survival (80%) in small gaps with tree shelters and landscape fabric. Differences between treatments were less pronounced in large gaps where survival was much lower (45%). In conclusion, seedlings are likely to have the greatest long-term survival if they are planted in smaller gaps protected from vegetative competition and rodent predation.

Multidimensional Measurement and Enhancement Strategies of the Provincial Ecological Niche in the Yellow River Basin

Provincial economic development in the VUCA era (V: volatility; U: uncertainty; C: complexity; A: ambiguity) is facing great challenges. Comprehensively measuring the provincial ecological niche and giving enhancement strategies are significant for the implementation of the Yellow River Strategy. By constructing the S–R–F (Status–Relationship–Function) provincial ecological niche measurement framework using the SEM model (the structural equation modeling), this paper calculated the provincial ecological niche of the Yellow River Basin from 2005 to 2019 using an ecological niche width model, an ecological niche overlap model and an ecological niche suitability model and gave improvement strategies through spatial positioning. Results: (1) The ecological niche breadth in the Yellow River Basin is uneven, showing a spatial pattern of “low in the upper reaches, high in the lower reaches”. The ecological niche overlap changes from synchronous competition to hierarchical differentiation. The ecological niche suitability shows a transformation of “overall homogeneity but local variation”. (2) Qinghai and Gansu belong to subspace I of ecological niches; the strategies of ecological niche specialization, potential ecological niche and ecological niche synergy are appropriate. Shanxi, Inner Mongolia and Ningxia belong to subspace III; the strategies of ecological niche specialization, ecological niche separation, potential ecological niche and ecological niche synergy are suitable. Shandong belongs to subspace VI; the strategies of ecological niche generalization and ecological niche synergy are fitting. Henan, Sichuan and Shaanxi belong to subspace VIII; it is advisable to choose ecological niche generalization, ecological niche separation and ecological niche synergy strategies. The possible marginal contributions of this paper are: (1) applying the idea of fitting optimization to guide the construction of a provincial ecological status indicator system, applying the validation analysis in the SEM model to test the suitability and validity of a provincial ecological status indicator system and improving the scientificity and objectivity of the indicator system construction. (2) Correlating the measurement model with the connotation (state–relationship–function) of the provincial ecological niche, the perspective of ecological niche research and improving the theoretical support for the construction of a multidimensional measurement model. (3) Spatial positioning of provincial ecological niches to improve the relevance and effectiveness of strategies.

Variation in the attached community structure and function of artificial habitats during ecological succession: A case study of the artificial reef area in Bailong Pearl Bay, China

With the continuous development of ports, ship transportation, resource extraction and artificial habitat deployment, an increasing number of artificial structures have created new usable spaces for marine organisms, and the biological communities living on their surfaces have undergone ecological succession. However, related studies have ignored ecological functions, interspecific relationships, and stability in different successional stages. Based on traditional taxonomy, this study applied functional diversity, niche overlap and niche breadth indexes to compare the attached organisms on artificial reefs (ARs) with different construction times in Bailong Pearl Bay, Fangchenggang City, Guangxi Zhuang Autonomous Region, China. The change trends of richness, habitat resource supply, resource utilization degree and efficiency, and interspecific relationships during the succession process were analysed. The results indicated that richness increased with functional diversity. While the number of species increased, the attached organisms showed different functional characteristics, and the number of species with identical traits decreased. The attached community in the later stage of development had a higher degree of utilization of ecological space, more comprehensive and efficient utilization of effective resources, a stronger degree of complementarity of organismal niches, and weaker interspecific competition. In the early stage of the developmental process, the attached community was affected by more environmental factors and had very significant correlations with those factors. The combination of the niche overlap index and the niche breadth index indicated that the resource supply supporting the survival and development of the attached organisms on the ARs with earlier construction times may be greater. The decrease in the number of biomarkers, the similarity of biomarkers belonging to the same family, and the similarity of dominance alternation all indicated that during the process of long-term development and community succession, the organisms attached to the various parts of the ARs tended to become more similar.

Effects of Spartina alterniflora invasion on the functional groups and niche of benthic crustaceans and fishes in mangrove wetland.

To investigate the effects of Spartina alterniflora invasion on mangrove wetland ecosystem, the benthic crustacean and fish samples were collected using cage nets in Zhangjiang Estuary Mangrove Wetland, Fujian Pro-vince in August 2020, January and April 2021. The five sampling sites included two mangroves (Avicennia marina and Kandelia obovata) sites, two S. alterniflora sites, and one mudflat site. The abundance, biomass, index of relative importance (IRI), Shannon diversity index (H), Pielou evenness index (J), and Margalef richness index (D) were used to quantify the dominant species and species diversity of benthic crustaceans and fishes. The functional groups and niche of dominant species were also analyzed. A total of 37 species, from 2 phyla, 2 classes, 8 orders, and 17 families, were identified across the three seasons. Most of them were warm-water and euryhaline species, mainly carnivorous and omnivorous functional groups. The results of two-way analysis of variance and NMDS showed that the community diversity index of benthic crustacean and fish did not change significantly after the invasion of S. alterniflora compared with mudflat. Functional groups had changed significantly, with the species number of plankton functional groups, carnivorous and phytophagous functional groups being increased. Compared with mangrove sites, the number of species of benthic crustacean and fish in S. alterniflora sites was significantly increased. The spatio-temporal niche breadth of dominant species ranged from 0 to 1.4186, with Liza carinata (1.4186), Bostrychus sinensis (1.0168), Metapenaeus ensis (0.9469) and Exopalaemon carinicauda (0.8922) as the top four species.

A dual involvement of Protocadherin 18a in stromal cell development guides the formation of a functional hematopoietic niche.

Hematopoietic stem and progenitor cells emerge from the aorta and migrate to the caudal hematopoietic tissue (CHT) of zebrafish larvae, the hematopoietic equivalent of the mammalian fetal liver, for their proliferation and differentiation. We previously reported that somite-derived stromal cells were a key component of the CHT niche. Here, we found that the cell adhesion protein Protocadherin 18a (Pcdh18a) is expressed in the stromal cell progenitors (SCPs) emigrating from somites toward the future CHT. Deletion of most of the Pcdh18a intracellular domain caused a decrease in the number of SCPs, the directionality of their migration, and the cell-contact mediated repulsion that normally occurs between migrating SCPs. These defects were followed by abnormal morphogenesis of the venous plexus that forms the CHT framework, and the inability of the CHT to function as a niche for hematopoietic stem and progenitor cells. Finally, we found that the extracellular domain of Pcdh18a mediates trans heterophilic adhesion of stromal cells to endothelial cells in vivo and thereby the reticular versus perivascular fate of SCPs. Thus, Pcdh18a expression in SCPs is essential for the proper development of the hematopoietic niche.

Two is better than one: Coupling DNA metabarcoding and stable isotope analysis improves dietary characterizations for a riparian-obligate, migratory songbird.

While an increasing number of studies are adopting molecular and chemical methods for dietary characterization, these studies often employ only one of these laboratory-based techniques; this approach may yield an incomplete, or even biased, understanding of diet due to each method's inherent limitations. To explore the utility of coupling molecular and chemical techniques for dietary characterizations, we applied DNA metabarcoding alongside stable isotope analysis to characterize the dietary niche of breeding Louisiana waterthrush (Parkesia motacilla), a migratory songbird hypothesized to preferentially provision its offspring with pollution-intolerant, aquatic arthropod prey. While DNA metabarcoding was unable to determine if waterthrush provision aquatic and terrestrial prey in different abundances, we found that specific aquatic taxa were more likely to be detected in successive seasons than their terrestrial counterparts, thus supporting the aquatic specialization hypothesis. Our isotopic analysis added greater context to this hypothesis by concluding that breeding waterthrush provisioned Ephemeroptera and Plecoptera, two pollution-intolerant, aquatic orders, in higher quantities than other prey groups, and expanded their functional trophic niche when such prey were not abundantly provisioned. Finally, we found that the dietary characterizations from each approach were often uncorrelated, indicating that the results gleaned from a diet study can be particularly sensitive to the applied methodologies. Our findings contribute to a growing body of work indicating the importance of high-quality, aquatic habitats for both consumers and their pollution-intolerant prey, while also demonstrating how the application of multiple, laboratory-based techniques can provide insights not offered by either technique alone.

Absolute quantification and genome-centric analyses elucidate the dynamics of microbial populations in anaerobic digesters

Anaerobic digestion (AD) relies on myriads of functions performed by complex microbial communities in customized settings, thus, a comprehensive investigation on the AD microbiome is central to the fine-tuned control. Most current AD microbiome studies are based on relative abundance, which hinders the interpretation of microbes’ dynamics and inter-sample comparisons. Here, we developed an absolute quantification (AQ) approach that integrated cellular spike-ins with metagenomic sequencing to elucidate microbial community variations and population dynamics in four anaerobic digesters. Using this method, 253 microbes were defined as decaying populations with decay rates ranging from -0.05 to -5.85 d−1, wherein, a population from Flavobacteriaceae family decayed at the highest rates of -3.87 to -5.85 d−1 in four digesters. Meanwhile, 25 microbes demonstrated the growing trend in the AD processes with growth rates ranging from 0.11 to 1.77 d−1, and genome-centric analysis assigned some of the populations to the functional niches of hydrolysis, short-chain fatty acids metabolism, and methane generation. Additionally, we observed that the specific activity of methanogens was lower in the prolonged digestion stage, and redundancy analysis revealed that the feedstock composition and the digestion duration were the two key parameters in governing the AD microbial compositions.

Bioengineering the Vascularized Endocrine Pancreas: A Fine-Tuned Interplay Between Vascularization, Extracellular-Matrix-Based Scaffold Architecture, and Insulin-Producing Cells.

Intrahepatic islet transplantation is a promising β-cell replacement strategy for the treatment of type 1 diabetes. Instant blood-mediated inflammatory reactions, acute inflammatory storm, and graft revascularization delay limit islet engraftment in the peri-transplant phase, hampering the success rate of the procedure. Growing evidence has demonstrated that islet engraftment efficiency may take advantage of several bioengineering approaches aimed to recreate both vascular and endocrine compartments either ex vivo or in vivo. To this end, endocrine pancreas bioengineering is an emerging field in β-cell replacement, which might provide endocrine cells with all the building blocks (vascularization, ECM composition, or micro/macro-architecture) useful for their successful engraftment and function in vivo. Studies on reshaping either the endocrine cellular composition or the islet microenvironment have been largely performed, focusing on a single building block element, without, however, grasping that their synergistic effect is indispensable for correct endocrine function. Herein, the review focuses on the minimum building blocks that an ideal vascularized endocrine scaffold should have to resemble the endocrine niche architecture, composition, and function to foster functional connections between the vascular and endocrine compartments. Additionally, this review highlights the possibility of designing bioengineered scaffolds integrating alternative endocrine sources to overcome donor organ shortages and the possibility of combining novel immune-preserving strategies for long-term graft function.

Competing constructions construct complementary niches

Abstract This paper traces the history of the English dative alternation by means of a quantitative analysis of instances of both the nominal and the prepositional construction in a corpus of Middle English (PPCME2), and compares the results to Wolk et al.’s (2013) data set from ARCHER. I show that the factors impacting the choice of one pattern over the other are subject to change over time: construction choice in Middle English is not straightforwardly predictable by the same factors at play in today’s alternation, but a clearer division based on syntactic semantic-pragmatic variables gradually emerges in the course to Late Modern English. I interpret this development as a prime case of competition, with a focus on (a) the initial emergence of functional overlap and thus competition, and (b) the subsequent creation of “functional niches” of the competing constructions.

Preface: Functional biogeography in Japanese cedar

Preface: Functional biogeography in Japanese cedar

Litter manipulation enhances plant community heterogeneity via distinct mechanisms: The role of distribution patterns of plant functional composition and niche breadth variability

Plant litter can greatly alter community compositional dynamics and variability of intraspecific interactions in grasslands, and thus the overall ecosystem structure and functions. However, whether plant activity can be driven by plant litter to modify plant community heterogeneity remains poorly explored. We investigate the responses of plant community heterogeneity to litter addition as well as their associated mechanisms. Here we conducted a three-year field experiment in a Tibetan alpine meadow to explore the effects of multiple plant litter addition (five mass levels and three species) on plant communities. We found that the effect of litter manipulation on plant community heterogeneity was mainly driven by litter mass rather than litter species. Higher litter mass manipulation significantly enhanced plant community heterogeneity, which was mainly determined by the niche breadth of forbs and the distribution patterns of functional composition rather than plant diversity. Our findings provide significant insights for understanding the effects of plant litter on grassland ecosystem dynamics to maintain the structure and function of ecosystems. Furthermore, this study suggests that reasonable management practices (e.g., moderate grazing in non−growing seasons) may be pivotal in achieving sustainability of grassland systems through plant litter dynamics.

Quantitative Analysis of Sympathetic and Nociceptive Innervation Across Bone Marrow Regions in Mice

Bone marrow (BM) innervation regulates the mobilization of hematopoietic stem and progenitor cells (HSPCs) from BM and stress hematopoiesis either by acting directly on HSPCs or by altering the niche function of mesenchymal and endothelial cells. However, the spatial distribution of BM innervation across bone regions is yet to be fully elucidated. Thus, we aimed to characterize the distribution of sympathetic and nociceptive nerves in each bone and BM region using three-dimensional quantitative microscopy. We discovered that sympathetic and nociceptive nerves were the major fibers throughout the BM. Compared with other femoral regions, central parts of the femoral BM were more densely innervated by both sympathetic and nociceptive nerves. Each region of the sternum was similarly innervated by sympathetic and nociceptive nerves. Further, the majority of sympathetic and nociceptive nerves in the BM ran parallel with arteries and arterioles, whereas the degree varied according to the bone type or BM region. In conclusion, this study provides spatial, topological, and functional information on BM innervation in a quantitative manner and illustrates that sympathetic and nociceptive nerves are two major components in BM innervation, mostly associated with arteries and arterioles.

Reconstructing the ecosystem context of a species: Honey-borne DNA reveals the roles of the honeybee.

To assess a species’ impact on its environment–and the environment’s impact upon a species–we need to pinpoint its links to surrounding taxa. The honeybee (Apis mellifera) provides a promising model system for such an exercise. While pollination is an important ecosystem service, recent studies suggest that honeybees can also provide disservices. Developing a comprehensive understanding of the full suite of services and disservices that honeybees provide is a key priority for such a ubiquitous species. In this perspective paper, we propose that the DNA contents of honey can be used to establish the honeybee’s functional niche, as reflected by ecosystem services and disservices. Drawing upon previously published genomic data, we analysed the DNA found within 43 honey samples from Northern Europe. Based on metagenomic analysis, we find that the taxonomic composition of DNA is dominated by a low pathogenicity bee virus with 40.2% of the reads, followed by bacteria (16.7%), plants (9.4%) and only 1.1% from fungi. In terms of ecological roles of taxa associated with the bees or taxa in their environment, bee gut microbes dominate the honey DNA, with plants as the second most abundant group. A range of pathogens associated with plants, bees and other animals occur frequently, but with lower relative read abundance, across the samples. The associations found here reflect a versatile the honeybee’s role in the North-European ecosystem. Feeding on nectar and pollen, the honeybee interacts with plants–in particular with cultivated crops. In doing so, the honeybee appears to disperse common pathogens of plants, pollinators and other animals, but also microbes potentially protective of these pathogens. Thus, honey-borne DNA helps us define the honeybee’s functional niche, offering directions to expound the benefits and drawbacks of the associations to the honeybee itself and its interacting organisms.

Development of micro-zymography: Visualization of enzymatic activity at the microscopic scale for aggregates collected from the rhizosphere

AimsVisualization of enzymatic activity links microbial functioning to localization in heterogeneous soil habitats. To assess enzymatic reactions in soil thin layer at the microscopic level, we developed a micro-zymography approach and tested it by visualization of the potential activity of phosphomonoesterase for aggregates collected from the rhizosphere of Zea mays L.MethodsWe evaluated micro-zymography by applying fluorogenically-labeled substrate i) on individual soil aggregates freshly sampled from the rhizosphere, ii) on thin layers of aggregates (≈ 500 µm) saturated with substrate to assess the dynamics of phosphomonoesterase activity, and iii) on maize roots under laser scanning microscope upon the identified hotspots by membrane-based zymography.ResultsWe found super transparent silicon as the most appropriate fixative to prevent sample drying. We optimized microscope settings to eliminate the soil auto-fluorescence. The fluorescent signal shifted from the free liquid phase towards the aggregate boundaries within 30 min after substrate addition and was finally detectable at the surface of a few aggregates. This was probably due to higher microbial abundance and enzymatic activity on the soil aggregates compared to the liquid phase. The enzymatic activity appeared patchy at the aggregate and root surfaces indicating heterogeneous distribution of hotspots.ConclusionsThe methodology including calibration, sample preparation, fixation, and monitoring was developed. The novel membrane-free micro-zymography approach is a promising tool to identify functional specificity and niche differentiation on roots and soil aggregates. This approach revealed unexplained complexity of competing processes (biochemical, hydrolytic, and physical) due to differently charged reaction products and enzyme-clay complexes.

Hedgehog signaling reprograms hair follicle niche fibroblasts to a hyper-activated state.

Hair follicle stem cells are regulated by dermal papilla fibroblasts, their principal signaling niche. Overactivation of Hedgehog signaling in the niche dramatically accelerates hair growth and induces follicle multiplication in mice. On single-cell RNA sequencing, dermal papilla fibroblasts increase heterogeneity to include new Wnt5ahigh states. Transcriptionally, mutant fibroblasts activate regulatory networks for Gli1, Alx3, Ebf1, Hoxc8, Sox18, and Zfp239. These networks jointly upregulate secreted factors for multiple hair morphogenesis and hair-growth-related pathways. Among these is non-conventional TGF-β ligand Scube3. We show that in normal mouse skin, Scube3 is expressed only in dermal papillae of growing, but not in resting follicles. SCUBE3 protein microinjection is sufficient to induce new hair growth, and pharmacological TGF-β inhibition rescues mutant hair hyper-activation phenotype. Moreover, dermal-papilla-enriched expression of SCUBE3 and its growth-activating effect are partially conserved in human scalp hair follicles. Thus, Hedgehog regulates mesenchymal niche function in the hair follicle via SCUBE3/TGF-β mechanism.

Unmasking the mammalian SET domain-containing protein 4.

SET domain-containing protein 4 (SETD4) is a member of a unique class of protein lysine methyltransferases. Here, we introduce the basic features of SETD4 and summarize the key findings from recent studies with emphases on its roles in tissue development and tumorigenesis, and its methylation substrates. SETD4 is expressed in stem/progenitor cells. Ablation of Setd4+ cells impedes the repopulation of acinar cells after pancreatic injury. Setd4 deletion in mice promotes the recovery of radiation-induced bone marrow (BM) failure by boosting the function of BM niche, facilitates the generation of endothelial cells and neovascularization of capillary vessels in the heart, enhances the proliferation of BM mesenchymal stem cells and disrupts the TLR4 signaling in BM-derived macrophages. SETD4 expression is also associated with the maintenance of quiescent breast cancer stem cells. While mouse Setd4 knockout delays radiation-induced T-lymphoma formation, elevated SETD4 expression has been observed in some proliferative cancer cells and is associated with a pro-survival potential. Oncogenic fusions of SETD4 have also been identified in cancer, albeit rare. In addition, SETD4 methylates lysine-570 in the C-terminal globular domain of KU70, which enables KU70 translocation to cytoplasm to suppress apoptosis.

P1402: ALTERATIONS OF HUMAN BONE MARROW NICHE IN CLONAL HEMATOPOIESIS AND AML

Background: Acute myeloid leukemia (AML) is a heterogeneous disease, predominantly caused by acquisition of mutations in the hematopoietic stem and progenitor cells. Some of these mutations are acquired in a preleukemic state termed clonal hematopoiesis of indeterminate potential (CHIP). Recent studies show that alterations in the bone marrow niche can support progression and therapy resistance of myeloid neoplasia. To date, it is unclear when these niche alterations occur and how they contribute to disease progression. In our study, we aim to elucidate alterations in the bone marrow niche, in particular in mesenchymal stem cells (MSC), during the evolution from healthy to CHIP and further to AML. Aims: Primary goal of this project is to examine how hematopoietic and niche cells from different health status (healthy, CHIP and AML) interact with each other and to what extent the presence of clonal hematopoiesis and leukemia have an impact on function and structure of the bone marrow niche. Methods: We collected primary human bone marrow (BM) and peripheral blood (pB) from hematologically healthy individuals who underwent hip replacement surgery at the LMU University Hospital. BM samples from AML patients are provided by the Laboratory for Leukemia Diagnostics, LMU University Hospital. Individuals are assigned to non-CHIP or CHIP cohorts based on targeted mutation analyses on pB samples. To study transcriptional changes in the bone marrow compartments, non-CHIP, CHIP and AML BM samples were FACS-sorted for 10 hematopoietic and non-hematopoietic cell types, and further processed for bulkRNA sequencing. MSC from donor BM samples were isolated via plastic adherence over 10-14 days of in vitro culture. MSC were characterized regarding their surface marker expression (FACS), differentiation (osteogenic, adipogenic, chondrogenic) potential and metabolic activity (Seahorse XF). Results: So far, we have included 98 healthy individuals and 64 have undergone targeted sequencing, revealing 19 individuals (30%) with CHIP. While there was no difference between the gender of the non-CHIP and CHIP groups, individuals with CHIP tended to be of older age (median, 64 y non-CHIP vs. 73.5 y CHIP). Most common mutations were DNMT3A (10/19), TET2 (7/19), ASXL1 (2/19), with variant allele frequencies varying between 1.0% and 16.8%, and 1.42 mutations per individual on average. The amount of mutations per individual increased with age. Analysis of MSCs from non-CHIP (n=5) and CHIP (n=5) individuals revealed no difference in surface marker expression over 10 passages. Differentiation assays of MSCs (non-CHIP=4, CHIP= 2) showed no difference in adipogenic and chondrogenic differentiation after 21 days. For osteogenesis, we noticed a trend towards an increased differentiation potential in the MSCs of CHIP individuals. To gain deeper insights into the BM niche we sorted 4 non-hematopoietic and 6 hematopoietic cell populations from non-CHIP (n=3), CHIP (n=4) and AML (n=4) individuals and performed bulkRNA sequencing on each population. So far changes in the composition of the bone marrow niche were only detected in the AML samples. Summary/Conclusion: Currently, we are analyzing the bulkRNA sequencing data and performing further experiments on cultured MSC to increase group sizes, as well as MSC and HSPC co-cultures in vitro to gain more insight into the bone marrow niche in non-CHIP, CHIP and AML. Additionally, we are continuously recruiting new individuals into our study. A comprehensive update on our cohort, sequencing data and MSC characterization will be shown at the conference.

Matrix-bound Cyr61/CCN1 is required to retain the properties of the bone marrow mesenchymal stem cell niche but is depleted with aging

Previously, we showed that extracellular matrices (ECMs), produced ex vivo by various types of stromal cells, direct bone marrow mesenchymal stem cells (BM-MSCs) in a tissue-specific manner and recapitulate physiologic changes characteristic of the aging microenvironment. In particular, BM-MSCs obtained from elderly donors and cultured on ECM produced by young BM stromal cells showed improved quantity, quality and osteogenic differentiation. In the present study, we searched for matrix components that are required for a functional BM-MSC niche by comparing ECMs produced by BM stromal cells from “young” (≤25 y/o) versus “elderly” (≥60 y/o) donors. With increasing donor age, ECM fibrillar organization and mechanical integrity deteriorated, along with the ability to promote BM-MSC proliferation and responsiveness to growth factors. Proteomic analyses revealed that the matricellular protein, Cyr61/CCN1, was present in young, but undetectable in elderly, BM-ECM. To assess the role of Cyr61 in the BM-MSC niche, we used genetic methods to down-regulate the incorporation of Cyr61 during production of young ECM and up-regulate its incorporation in elderly ECM. The results showed that Cyr61-depleted young ECM lost the ability to promote BM-MSC proliferation and growth factor responsiveness. However, up-regulating the incorporation of Cyr61 during synthesis of elderly ECM restored its ability to support BM-MSC responsiveness to osteogenic factors such as BMP-2 and IGF-1. We next examined aging bone and compared bone mineral density and Cyr61 content of L4-L5 vertebral bodies in “young” (9–11 m/o) and “elderly” (21–33 m/o) mice. Our analyses showed that low bone mineral density was associated with decreased amounts of Cyr61 in osseous tissue of elderly versus young mice. Our results strongly demonstrate a novel role for ECM-bound Cyr61 in the BM-MSC niche, where it is responsible for retention of BM-MSC proliferation and growth factor responsiveness, while depletion of Cyr61 from the BM niche contributes to an aging-related dysregulation of BM-MSCs. Our results also suggest new potential therapeutic targets for treating age-related bone loss by restoring specific ECM components to the stem cell niche.

S245: ALTERATION OF BONE MARROW NICHE BY ALLOGENEIC IMMUNE REACTION AFTER HSCT

Background: Allogeneic stem cell transplantation (allo-SCT) has been considered as a curable therapy for hematologic diseases. Post-transplant myelosuppression is one of the life-threatening complications. Hematopoietic stem cell (HSC) functions are tightly regulated by a specialized microenvironment called “niche” in the bone marrow (BM) and we have identified differential contributions of perivascular stromal cells as HSC niche (Asada et al. Nat Cell Biol 2017). Recent studies indicate that BM niche can be targeted by allo-immune reaction but it remains unclear how the perivascular niche cells are involved in the mechanism of HSC dysfunction after allo-SCT. Aims: In this study, we aim to investigate the mechanisms of the impairment of the niche cells by allo-immune reactions, and to explore a targetable pathway for myelosuppression after allo-SCT. Methods: To evaluate the alterations in niche cells after allo-SCT, we utilized allo-SCT mice models (C3H/HeJ into C57BL/6, C57BL/6 into B6D2F1). Hematopoietic cells, including HSCs and mature cells, and perivascular stromal cells in BM were analyzed by flow cytometry (FACS) after transplantation. In imaging studies, we visualized perivascular stromal cells using Nestin-GFP (Nes-GFP), Myh11-CreER/tdTomato, LeptinR-Cre/tdTomato transgenic mice and observed them with confocal laser scanning microscopy. The gene expression analysis of niche factors was performed by real-time PCR. To assess the landscape of transcriptional remodeling in BM after allo-SCT, we performed single-cell RNA sequencing (scRNAseq) of BM cells. Results: FACS analyses showed that Nes-GFP+ stromal cells were dramatically reduced, and HSC recovery was severely impaired in allo-SCT mice compared to control mice at day 21 after SCT. Imaging studies also revealed that perivascular stromal cells are morphologically damaged in allo-SCT mice. Nes-GFP+ stromal cells in allo-SCT mice presented a decreased expression of niche factors, including Cxcl12 and Scf, which are essential for HSC maintenance, indicating the dysfunction of niche cells. We confirmed the impaired niche function in allo-SCT mice by tandem transplantation analysis in vivo, in which the recovery of syngeneic HSCs was also impaired in damaged microenvironment in allo-SCT mice. Prevention of allo-reactive T cell expansion by post-transplant cyclophosphamide ameliorated the niche cell reduction along with improved recovery of HSCs. These results indicate that niche impairment caused by allo-immune reaction leads to HSC dysfunction. The scRNAseq of BM highlighted the elevated expression levels of inflammation response genes, including Cxcl9 and Cxcl10, and MHC classⅡ, together with the deceased expression of niche factors in niche cells in allo-SCT mice. These alterations seemed to be induced by proinflammatory cytokines such as interferon-gamma (INF-γ) derived from allo-T cells, as the increased number of T cells and high levels of INF-γ in BM were observed in allo-SCT mice. Additionally, evaluation of T cell localization by using imaging techniques of BM revealed that allo-T cells reside significant closer to LepR+ niche cells in allo-SCT mice. Since activation of CXCR3, which is a receptor for chemokines Cxcl9 and Cxcl10, has been shown to induce chemotaxis of activated T cells, we treated allo-SCT mice with AMG487, an antagonist of CXCR3, resulted in an improved recovery of both perivascular niche cells and hematopoiesis in allo-SCT mice. Summary/Conclusion: Collectively, allo-immune reaction severely damages perivascular HSC niche cells, leading to impaired recovery of hematopoiesis after allo-SCT.

Contrasting adaptation and optimization of stomatal traits across communities at continental scale.

Shifts in stomatal trait distributions across contrasting environments and their linkage with ecosystem productivity at large spatial scales have been unclear. Here, we measured the maximum stomatal conductance (g), stomatal area fraction (f), and stomatal space-use efficiency (e, the ratio of g to f) of 800 plant species ranging from tropical to cold-temperate forests, and determined their values for community-weighted mean, variance, skewness, and kurtosis. We found that the community-weighted means of g and f were higher in drier sites, and thus, that drought 'avoidance' by water availability-driven growth pulses was the dominant mode of adaptation for communities at sites with low water availability. Additionally, the variance of g and f was also higher at arid sites, indicating greater functional niche differentiation, whereas that for e was lower, indicating the convergence in efficiency. When all other stomatal trait distributions were held constant, increasing kurtosis or decreasing skewness of g would improve ecosystem productivity, whereas f showed the opposite patterns, suggesting that the distributions of inter-related traits can play contrasting roles in regulating ecosystem productivity. These findings demonstrate the climatic trends of stomatal trait distributions and their significance in the prediction of ecosystem productivity.