Fate Mapping via Ms4a3-Expression History Traces Monocyte-Derived Cells.

Most tissue-resident macrophage (RTM) populations are seeded by waves of embryonic hematopoiesis and are self-maintained independently of a bone marrow contribution during adulthood. A proportion of RTMs, however, is constantly replaced by blood monocytes, and their functions compared to embryonic RTMs remain unclear. The kinetics and extent of the contribution of circulating monocytes to RTM replacement during homeostasis, inflammation, and disease are highly debated. Here, we identified Ms4a3 as a specific gene expressed by granulocyte-monocyte progenitors (GMPs) and subsequently generated Ms4a3TdT reporter, Ms4a3Cre, and Ms4a3CreERT2 fate-mapping models. These models traced efficiently monocytes and granulocytes, but no lymphocytes or tissue dendritic cells. Using these models, we precisely quantified the contribution of monocytes to the RTM pool during homeostasis and inflammation. The unambiguous identification of monocyte-derived cells will permit future studies of their function under any condition.

Author Correction: Requirement for the basic helix-loop-helix transcription factor Dec2 in initial TH2 lineage commitment.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

MAP3K2-regulated intestinal stromal cells define a distinct stem cell niche.

Intestinal stromal cells are known to modulate the propagation and differentiation of intestinal stem cells1,2. However, the precise cellular and molecular mechanisms by which this diverse stromal cell population maintains tissue homeostasis and repair are poorly understood. Here we describe a subset of intestinal stromal cells, named MAP3K2-regulated intestinal stromal cells (MRISCs), and show that they are the primary cellular source of the WNT agonist R-spondin 1 following intestinal injury in mice. MRISCs, which are epigenetically and transcriptomically distinct from subsets of intestinal stromal cells that have previously been reported3-6, are strategically localized at the bases of colon crypts, and function to maintain LGR5+ intestinal stem cells and protect against acute intestinal damage through enhanced R-spondin 1 production. Mechanistically, this MAP3K2 specific function is mediated by a previously unknown reactive oxygen species (ROS)-MAP3K2-ERK5-KLF2 axis to enhance production of R-spondin 1. Our results identify MRISCs as a key component of an intestinal stem cell niche that specifically depends on MAP3K2 to augment WNT signalling for the regeneration of damaged intestine.

A Tunable Diffusion-Consumption Mechanism of Cytokine Propagation Enables Plasticity in Cell-to-Cell Communication in the Immune System.

Immune cells communicate by exchanging cytokines to achieve a context-appropriate response, but the distances over which such communication happens are not known. Here, we used theoretical considerations and experimental models of immune responses in vitro and in vivo to quantify the spatial extent of cytokine communications in dense tissues. We established that competition between cytokine diffusion and consumption generated spatial niches of high cytokine concentrations with sharp boundaries. The size of these self-assembled niches scaled with the density of cytokine-consuming cells, a parameter that gets tuned during immune responses. In vivo, we measured interactions on length scales of 80-120 µm, which resulted in a high degree of cell-to-cell variance in cytokine exposure. Such heterogeneous distributions of cytokines were a source of non-genetic cell-to-cell variability that is often overlooked in single-cell studies. Our findings thus provide a basis for understanding variability in the patterning of immune responses by diffusible factors.

PLK4 deubiquitination by Spata2-CYLD suppresses NEK7-mediated NLRP3 inflammasome activation at the centrosome.

The innate immune sensor NLRP3 assembles an inflammasome complex with NEK7 and ASC to activate caspase-1 and drive the maturation of proinflammatory cytokines IL-1ß and IL-18. NLRP3 inflammasome activity must be tightly controlled, as its over-activation is involved in the pathogenesis of inflammatory diseases. Here, we show that NLRP3 inflammasome activation is suppressed by a centrosomal protein Spata2. Spata2 deficiency enhances NLRP3 inflammasome activity both in the macrophages and in an animal model of peritonitis. Mechanistically, Spata2 recruits the deubiquitinase CYLD to the centrosome for deubiquitination of polo-like kinase 4 (PLK4), the master regulator of centrosome duplication. Deubiquitination of PLK4 facilitates its binding to and phosphorylation of NEK7 at Ser204. NEK7 phosphorylation in turn attenuates NEK7 and NLRP3 interaction, which is required for NLRP3 inflammasome activation. Pharmacological or shRNA-mediated inhibition of PLK4, or mutation of the NEK7 Ser204 phosphorylation site, augments NEK7 interaction with NLRP3 and causes increased NLRP3 inflammasome activation. Our study unravels a novel centrosomal regulatory pathway of inflammasome activation and may provide new therapeutic targets for the treatment of NLRP3-associated inflammatory diseases.

An Ultrasensitive Ti3C2Tx MXene-based Soft Contact Lens for Continuous and Nondestructive Intraocular Pressure Monitoring.

Wearable soft contact lens sensors for continuous and nondestructive intraocular pressure (IOP) monitoring are highly desired as glaucoma and postoperative myopia patients grow, especially as the eyestrain crowd increases. Herein, a smart closed-loop system is presented that combines a Ti3C2Tx MXene-based soft contact lens (MX-CLS) sensor, wireless data transmission units, display, and warning components to realize continuous and nondestructive IOP monitoring/real-time display. The fabricated MX-CLS device exhibits an extremely high sensitivity of 7.483 mV mmHg-1, good linearity on silicone eyeballs, excellent stability under long-term pressure-release measurement, sufficient transparency with 67.8% transmittance under visible illumination, and superior biocompatibility with no discomfort when putting the MX-CLS sensor onto the Rabbit eyes. After integrating with the wireless module, users can realize real-time monitoring and warning of IOP via smartphones, the demonstrated MX-CLS device together with the IOP monitoring/display system opens up promising platforms for Ti3C2Tx materials as the base for multifunctional contact lens-based sensors and continuous and nondestructive IOP measurement system.

CD169+ Skin Macrophages Function as a Specialized Subpopulation in Promoting Psoriasis-like Skin Disease in Mice.

Skin macrophages are critical to maintain and restore skin homeostasis. They serve as major producers of cytokines and chemokines in the skin, participating in diverse biological processes such as wound healing and psoriasis. The heterogeneity and functional diversity of macrophage subpopulations endow them with multifaceted roles in psoriasis development. A distinct subpopulation of skin macrophages, characterized by high expression of CD169, has been reported to exist in both mouse and human skin. However, its role in psoriasis remains unknown. Here, we report that CD169+ macrophages exhibit increased abundance in imiquimod (IMQ) induced psoriasis-like skin lesions. Specific depletion of CD169+ macrophages in CD169-ditheria toxin receptor (CD169-DTR) mice inhibits IMQ-induced psoriasis, resulting in milder symptoms, diminished proinflammatory cytokine levels and reduced proportion of Th17 cells within the skin lesions. Furthermore, transcriptomic analysis uncovers enhanced activity in CD169+ macrophages when compared with CD169- macrophages, characterized by upregulated genes that are associated with cell activation and cell metabolism. Mechanistically, CD169+ macrophages isolated from IMQ-induced skin lesions produce more proinflammatory cytokines and exhibit enhanced ability to promote Th17 cell differentiation in vitro. Collectively, our findings highlight the crucial involvement of CD169+ macrophages in psoriasis development and offer novel insights into the heterogeneity of skin macrophages in the context of psoriasis.

CD169+ subcapsular sinus macrophage-derived microvesicles are associated with light zone follicular dendritic cells.

Follicular dendritic cells (FDCs) are a specialized type of stromal cells that exclusively reside in B-cell follicles. When inflammation occurs, the FDC network is reorganized to support germinal center (GC) polarization into the light zone (LZ) and dark zone (DZ). Despite the indispensable role of FDCs in supporting humoral responses, the FDC regulatory requirements remain incompletely defined. In this study, we unexpectedly observed an accumulation of CD169+ subcapsular sinus macrophage (SSM)-derived microvesicles (MVs) in the B-cell zone, which were tightly associated with the FDC network. Interestingly, a selective deposition of CD169+ MVs was detected in both GC LZ FDCs in secondary follicles and on predetermined LZ FDCs in primary follicles. The ablation of CD169+ MVs, resulting from SSM depletion, resulted in significantly decreased expression of LZ-related genes in FDCs. In addition, we found that CD169+ MVs could colocalize with fluorescently tagged antigen-containing immune complexes (ICs), supporting a possible role of CD169+ MVs in transporting antigens to the FDC network. Thus, our data reveal intimate crosstalk between FDCs and SSMs located outside B-cell follicles via SSM-released MVs, providing a novel perspective on the mechanisms underlying the regulation of FDC maturation and polarization.

ECM1 controls T(H)2 cell egress from lymph nodes through re-expression of S1P(1).

Type 2 helper T cells (T(H)2) are critically involved in allergies and asthma. Here we demonstrate that extracellular matrix protein-1 (ECM1) is highly and selectively expressed in T(H)2 cells. ECM1 deficiency caused impaired T(H)2 responses and reduced allergic airway inflammation in vivo. Functional analysis demonstrated that although the T(H)2 polarization of ECM1-deficient cells was unimpaired, these cells had a defect in migration and were retained in peripheral lymphoid organs. This was associated with reduced expression of KLF2 and S1P(1). We also found that ECM1 could directly bind the interleukin-2 (IL-2) receptor to inhibit IL-2 signaling and activate S1P(1) expression. Our data identify a previously unknown function of ECM1 in regulating T(H)2 cell migration through control of KLF2 and S1P(1) expression.

Requirement for the basic helix-loop-helix transcription factor Dec2 in initial TH2 lineage commitment.

How naive CD4(+) T cells commit to the T helper type 2 (T(H)2) lineage is poorly understood. Here we show that the basic helix-loop-helix transcription factor Dec2 was selectively expressed in T(H)2 cells. CD4(+) T cells from Dec2-deficient mice showed defective T(H)2 differentiation in vitro and in vivo in an asthma model and in response to challenge with a parasite antigen. Dec2 promoted expression of interleukin 4 (IL-4), IL-5 and IL-13 during early T(H)2 differentiation and directly bound to and activated transcription of genes encoding the transcription factors JunB and GATA-3. As GATA-3 induces Dec2 expression, our findings also indicate a feed-forward regulatory circuit during T(H)2 differentiation.

Immune homeostasis enforced by co-localized effector and regulatory T cells.

FOXP3(+) regulatory T cells (Treg cells) prevent autoimmunity by limiting the effector activity of T cells that have escaped thymic negative selection or peripheral inactivation. Despite the information available about molecular factors mediating the suppressive function of Treg cells, the relevant cellular events in intact tissues remain largely unexplored, and whether Treg cells prevent activation of self-specific T cells or primarily limit damage from such cells has not been determined. Here we use multiplex, quantitative imaging in mice to show that, within secondary lymphoid tissues, highly suppressive Treg cells expressing phosphorylated STAT5 exist in discrete clusters with rare IL-2-positive T cells that are activated by self-antigens. This local IL-2 induction of STAT5 phosphorylation in Treg cells is part of a feedback circuit that limits further autoimmune responses. Inducible ablation of T cell receptor expression by Treg cells reduces their regulatory capacity and disrupts their localization in clusters, resulting in uncontrolled effector T cell responses. Our data thus reveal that autoreactive T cells are activated to cytokine production on a regular basis, with physically co-clustering T cell receptor-stimulated Treg cells responding in a negative feedback manner to suppress incipient autoimmunity and maintain immune homeostasis.

Promising Fast Energy Transfer System Between Graphene Quantum Dots and the Application in Fluorescent Bioimaging.

Tunable photoluminescence performance of graphene quantum dots (GQDs) is one of the most important topics for the development of GQDs. In this paper, we report lattice-doped GQDs (boron-doped GQDs (B-GQDs) and phosphorus-doped GQDs (P-GQDs)). Because of the matched band structure, the fast energy transfer between blue-emitted B-GQDs (emission wavelength: 460 nm) and orange-emitted P-GQDs (emission wavelength: 630 nm) can induce an efficient fluorescence emission in P-GQDs once B-GQDs are excited under the optimal excitation wavelength of 460 nm. Moreover, with the effective energy transfer, the quantum yield of P-GQDs increased to 0.48, which is much higher than that of pure P-GQDs. We also demonstrated the potentials of this system for fluorescent bioimaging in vitro.

Allergen-Induced CD4+ T Cell Cytokine Production within Airway Mucosal Dendritic Cell-T Cell Clusters Drives the Local Recruitment of Myeloid Effector Cells.

Allergic asthma develops in the mucosal tissue of small bronchi. At these sites, local cytokine production by Th2/Th17 cells is believed to be critical for the development of tissue eosinophilia/neutrophilia. Using the mouse trachea as a relevant model of human small airways, we performed advanced in vivo dynamic and in situ static imaging to visualize individual cytokine-producing T cells in the airway mucosa and to define their immediate cellular environment. Upon allergen sensitization, newly recruited CD4+ T cells formed discrete Ag-driven clusters with dendritic cells (DCs). Within T cell-DC clusters, a small fraction of CD4+ T cells produced IL-13 or IL-17 following prolonged Ag-specific interactions with DCs. As a result of local Th2 cytokine signaling, eosinophils were recruited into these clusters. Neutrophils also infiltrated these clusters in a T cell-dependent manner, but their mucosal distribution was more diffuse. Our findings reveal the focal nature of allergen-driven responses in the airways and define multiple steps with potential for interference with the progression of asthmatic pathology.

The Synergistic Effect on the Mimetic Optical Structure of Feline Eyes toward Household Health Monitoring of Acute and Chronic Diseases.

A breakthrough in the performance of bionic optical structures will only be achieved if we can obtain an in-depth understanding of the synergy mechanisms operating in natural optical structures and find ways to imitate them. In this work, inspired by feline eyes, an optical substrate that takes advantage of a synergistic effect that occurs between resonant and reflective structures was designed. The synergistic effect between the reflective and resonant components leads to a Raman enhancement factor (EF) of 1.16 × 107, which is much greater than that achieved using the reflective/resonant cavities on their own. Finite-difference time-domain (FDTD) simulations and experimental results together confirm that the mechanism of this synergistic effect is achieved by realizing multiple reflections and repeated absorptions of light, generating a strong local electric field. Thus, a 2-3 order of magnitude increase in sensitivity could be achieved. More importantly, with the homemade centrifugal device, above optical substrates were further used to develop a rapidly highly sensitive household health monitoring system (detection time <3 min). It can thus be used to give early warning of acute diseases with high risk (e.g., acute myocardial infarction (AMI) and cerebral peduncle). Due to the good reusability and storability (9% and 8% reduction in EF after washing 30 times and 9 months of storage, respectively) of the substrates, the substrates thus reduce detection costs (to ∼$1), making them much cheaper to use than the current gold-standard methods (e.g., ∼$16 for gout detection).

Hypothalamic warm-sensitive neurons require TRPC4 channel for detecting internal warmth and regulating body temperature in mice.

Precise monitoring of internal temperature is vital for thermal homeostasis in mammals. For decades, warm-sensitive neurons (WSNs) within the preoptic area (POA) were thought to sense internal warmth, using this information as feedback to regulate body temperature (Tcore). However, the cellular and molecular mechanisms by which WSNs measure temperature remain largely undefined. Via a pilot genetic screen, we found that silencing the TRPC4 channel in mice substantially attenuated hypothermia induced by light-mediated heating of the POA. Loss-of-function studies of TRPC4 confirmed its role in warm sensing in GABAergic WSNs, causing additional defects in basal temperature setting, warm defense, and fever responses. Furthermore, TRPC4 antagonists and agonists bidirectionally regulated Tcore. Thus, our data indicate that TRPC4 is essential for sensing internal warmth and that TRPC4-expressing GABAergic WSNs function as a novel cellular sensor for preventing Tcore from exceeding set-point temperatures. TRPC4 may represent a potential therapeutic target for managing Tcore.

Distributed implantation of a flexible microelectrode array for neural recording.

Flexible multichannel electrode arrays (fMEAs) with multiple filaments can be flexibly implanted in various patterns. It is necessary to develop a method for implanting the fMEA in different locations and at various depths based on the recording demands. This study proposed a strategy for reducing the microelectrode volume with integrated packaging. An implantation system was developed specifically for semiautomatic distributed implantation. The feasibility and convenience of the fMEA and implantation platform were verified in rodents. The acute and chronic recording results provied the effectiveness of the packaging and implantation methods. These methods could provide a novel strategy for developing fMEAs with more filaments and recording sites to measure functional interactions across multiple brain regions.

Investigation of a Highly Sensitive Surface-Enhanced Raman Scattering Substrate Formed by a Three-Dimensional/Two-Dimensional Graphene/Germanium Heterostructure.

Three-dimensional graphene (3D-graphene) is used in surface-enhanced Raman spectroscopy (SERS) because of its plasmonic nanoresonator structure and good ability to interact with light. However, a thin (3-5 nm) layer of amorphous carbon (AC) inevitably appears as a template layer between the 3D-graphene and object substrate when the 3D-graphene layer is synthesized, weakening the enhancement factor. Herein, two-dimensional graphene (2D-graphene) is employed as a template layer to directly synthesize 3D-graphene on a germanium (Ge) substrate via plasma-assisted chemical vapor deposition, bypassing the formation of an AC layer. The interaction and photoinduced charge transfer ability of the 3D-graphene/Ge heterojunction with incident light are improved. Moreover, the high density of electronic states close to the Fermi level of the heterojunction induces the adsorbed probe molecules to efficiently couple to the 3D-graphene-based SERS substrate. Our experimental results imply that the lowest concentrations of rhodamine 6G and rhodamine B that can be detected on the 3D/2D-graphene/Ge SERS substrate correspond to 10-10 M; for methylene blue, it is 10-8 M. The detection limits of the 3D/2D-graphene/Ge SERS substrate with respect to 3-hydroxytyramine hydrochloride and melamine (in milk) are both less than 1 ppm. This work may provide a viable and convenient alternative method for preparing 3D-graphene SERS substrates. It also constitutes a new approach to developing SERS substrates with remarkable performance levels.

Dec2 promotes Th2 cell differentiation by enhancing IL-2R signaling.

Th cell differentiation is precisely regulated by thousands of genes at different stages. In the present study, we demonstrate that Dec2, a transcription factor belonging to the bHLH (basic helix-loop-helix) superfamily, is progressively induced during the course of Th2 differentiation, especially at the late stage. The up-regulated Dec2 can strongly promote Th2 development under Th2-inducing conditions, as evidenced by retrovirus-mediated gene transfer or transgenic manipulation. In addition, an enhancement of Th2 responses is also detectable in Dec2 transgenic mice in vivo. Conversely, RNA interference-mediated suppression of endogenous Dec2 could attenuate Th2 differentiation. Finally, we show that the enhanced Th2 development is at least in part due to substantial up-regulation of CD25 expression elicited by Dec2, thereby resulting in hyperresponsiveness to IL-2 stimulation.

Wearable Contact Lens Sensor for Non-invasive Continuous Monitoring of Intraocular Pressure.

Intraocular pressure (IOP) is an essential indicator of the diagnosis and treatment of glaucoma. IOP has an apparent physiological rhythm, and it often reaches its peak value at night. To avoid missing the peak value at night and sample the entire rhythm cycle, the continuous monitoring of IOP is urgently needed. A wearable contact lens IOP sensor based on a platinum (Pt) strain gauge is fabricated by the micro-electro-mechanical (MEMS) process. The structure and parameters of the strain gauge are optimized to improve the sensitivity and temperature stability. Tests on an eyeball model indicate that the IOP sensor has a high sensitivity of 289.5 µV/mmHg and excellent dynamic cycling performance at different speeds of IOP variation. The temperature drift coefficient of the sensor is 33.4 µV/°C. The non-invasive IOP sensor proposed in this report exhibits high sensitivity and satisfactory stability, promising a potential in continuous IOP monitoring.

Sensitive, Reusable, Surface-Enhanced Raman Scattering Sensors Constructed with a 3D Graphene/Si Hybrid.

Surface-enhanced Raman scattering (SERS) substrates based on graphene and its derivatives have recently attracted attention among those interested in the detection of trace molecules; however, these substrates generally show poor uniformity, an unsatisfactory enhancement factor, and require a complex fabrication process. Herein, we design and fabricate three-dimensional (3D) graphene/silicon (3D-Gr/Si) heterojunction SERS substrates to detect various types of molecules. Notably, the detection limit of 3D-Gr/Si can reach 10-10 M for rhodamine 6G (R6G) and rhodamine B (RB), 10-7 M for crystal violet (CRV), copper(II) phthalocyanine (CuPc), and methylene blue (MB), 10-8 M for dopamine (DA), 10-6 M for bovine serum albumin (BSA), and 10-5 M for melamine (Mel), which is superior to most reported graphene-based SERS substrates. Besides, the proposed 3D-Gr/Si heterojunction SERS substrates can achieve a high uniformity with relative standard deviations (RSDs) of less than 5%. Moreover, the 3D-Gr/Si SERS substrates are reusable after washing with ethyl alcohol to remove the adsorbed molecules. These excellent SERS performances are attributed to the novel 3D structure and abundantly exposed atomically thin edges, which facilitate charge transfer between 3D-Gr and probe molecules. We believe that the 3D-Gr/Si heterojunction SERS substrates offer potential for practical applications in biochemical molecule detection and provide insight into the design of high-performance SERS substrates.