Aged microglia promote peripheral T cell infiltration by reprogramming the microenvironment of neurogenic niches.

BACKGROUND: The immune cell compartment of the mammalian brain changes dramatically and peripheral T cells infiltrate the brain parenchyma during normal aging. However, the mechanisms underlying age-related T cell infiltration in the central nervous system remain unclear. RESULTS: Chronic inflammation and peripheral T cell infiltration were observed in the subventricular zone of aged mice. Cell-cell interaction analysis revealed that aged microglia released CCL3 to recruit peripheral CD8+ memory T cells. Moreover, the aged microglia shifted towards a pro-inflammation state and released TNF-α to upregulate the expression of VCAM1 and ICAM1 in brain venous endothelial cells, which promoted the transendothelial migration of peripheral T cells. In vitro experiment reveals that human microglia would also transit to a chemotactic phenotype when treated with CSF from the elderly. CONCLUSIONS: Our research demonstrated that microglia play an important role in the aging process of brain by shifting towards a pro-inflammation and chemotactic state. Aged microglia promote T cell infiltration by releasing chemokines and upregulating adhesion molecules on venous brain endothelial cells.

Mer regulates microglial/macrophage M1/M2 polarization and alleviates neuroinflammation following traumatic brain injury.

BACKGROUND: Traumatic brain injury (TBI) is a leading cause of death and disability worldwide. Microglial/macrophage activation and neuroinflammation are key cellular events following TBI, but the regulatory and functional mechanisms are still not well understood. Myeloid-epithelial-reproductive tyrosine kinase (Mer), a member of the Tyro-Axl-Mer (TAM) family of receptor tyrosine kinases, regulates multiple features of microglial/macrophage physiology. However, its function in regulating the innate immune response and microglial/macrophage M1/M2 polarization in TBI has not been addressed. The present study aimed to evaluate the role of Mer in regulating microglial/macrophage M1/M2 polarization and neuroinflammation following TBI. METHODS: The controlled cortical impact (CCI) mouse model was employed. Mer siRNA was intracerebroventricularly administered, and recombinant protein S (PS) was intravenously applied for intervention. The neurobehavioral assessments, RT-PCR, Western blot, magnetic-activated cell sorting, immunohistochemistry and confocal microscopy analysis, Nissl and Fluoro-Jade B staining, brain water content measurement, and contusion volume assessment were performed. RESULTS: Mer is upregulated and regulates microglial/macrophage M1/M2 polarization and neuroinflammation in the acute stage of TBI. Mechanistically, Mer activates the signal transducer and activator of transcription 1 (STAT1)/suppressor of cytokine signaling 1/3 (SOCS1/3) pathway. Inhibition of Mer markedly decreases microglial/macrophage M2-like polarization while increases M1-like polarization, which exacerbates the secondary brain damage and sensorimotor deficits after TBI. Recombinant PS exerts beneficial effects in TBI mice through Mer activation. CONCLUSIONS: Mer is an important regulator of microglial/macrophage M1/M2 polarization and neuroinflammation, and may be considered as a potential target for therapeutic intervention in TBI.

Ceria nanoparticles ameliorate white matter injury after intracerebral hemorrhage: microglia-astrocyte involvement in remyelination.

BACKGROUND: Intracerebral hemorrhage (ICH) can induce excessive accumulation of reactive oxygen species (ROS) that may subsequently cause severe white matter injury. The process of oligodendrocyte progenitor cell (OPC) differentiation is orchestrated by microglia and astrocytes, and ROS also drives the activation of microglia and astrocytes. In light of the potent ROS scavenging capacity of ceria nanoparticles (CeNP), we aimed to investigate whether treatment with CeNP ameliorates white matter injury by modulating ROS-induced microglial polarization and astrocyte alteration. METHODS: ICH was induced in vivo by collagenase VII injection. Mice were administered with PLX3397 for depleting microglia. Primary microglia and astrocytes were used for in vitro experiments. Transmission electron microscopy analysis and immunostaining were performed to verify the positive effects of CeNP in remyelination and OPC differentiation. Flow cytometry, real-time polymerase chain reaction, immunofluorescence and western blotting were used to detect microglia polarization, astrocyte alteration, and the underlying molecular mechanisms. RESULTS: CeNP treatment strongly inhibited ROS-induced NF-κB p65 translocation in both microglia and astrocytes, and significantly decreased the expression of M1 microglia and A1 astrocyte. Furthermore, we found that CeNP treatment promoted remyelination and OPC differentiation after ICH, and such effects were alleviated after microglial depletion. Interestingly, we also found that the number of mature oligodendrocytes was moderately increased in ICH + CeNP + PLX3397-treated mice compared to the ICH + vehicle + PLX3397 group. Therefore, astrocytes might participate in the pathophysiological process. The subsequent phagocytosis assay indicated that A1 astrocyte highly expressed C3, which could bind with microglia C3aR and hinder microglial engulfment of myelin debris. This result further replenished the feedback mechanism from astrocytes to microglia. CONCLUSION: The present study reveals a new mechanism in white matter injury after ICH: ICH induces M1 microglia and A1 astrocyte through ROS-induced NF-κB p65 translocation that hinders OPC maturation. Subsequently, A1 astrocytes inhibit microglial phagocytosis of myelin debris via an astrocytic C3-microglial C3aR axis. Polyethylene glycol-CeNP treatment inhibits this pathological process and ultimately promotes remyelination. Such findings enlighten us that astrocytes and microglia should be regarded as a functional unit in future works.

Construction of competitive endogenous RNA network reveals regulatory role of long non-coding RNAs in intracranial aneurysm.

BACKGROUND: Rupture of intracranial aneurysm (IA) is the main cause of devastating subarachnoid hemorrhage, which urges our understanding of the pathogenesis and regulatory mechanisms of IA. However, the regulatory roles of long non-coding RNAs (lncRNAs) in IA is less known. RESULTS: We processed the raw SRR files of 12 superficial temporal artery (STA) samples and 6 IA samples to count files. Then the differentially expressed (DE) mRNAs, miRNAs, and lncRNAs between STAs and IAs were identified. The enrichment analyses were performed using DEmRNAs. Next, a lncRNA-miRNA-mRNA regulatory network was constructed using integrated bioinformatics analysis. In summary, 341 DElncRNAs, 234 DEmiRNAs, and 2914 DEmRNAs between the STA and IA. The lncRNA-miRNA-mRNA regulatory network of IA contains 91 nodes and 146 edges. The subnetwork of hub lncRNA PVT1 was extracted. The expression level of PVT1 was positively correlated with a majority of the mRNAs in its subnetwork. Moreover, we found that several mRNAs (CCND1, HIF1A, E2F1, CDKN1A, VEGFA, COL1A1 and COL5A2) in the PVT1 subnetwork served as essential components in the PI3K-Akt signaling pathway, and that some of the non-coding RNAs (ncRNAs) (PVT1, HOTAIR, hsa-miR-17, hsa-miR-142, hsa-miR-383 and hsa-miR-193b) interacted with these mRNAs. CONCLUSION: Our annotations noting ncRNA's role in the pathway may uncover novel regulatory mechanisms of ncRNAs and mRNAs in IA. These findings provide significant insights into the lncRNA regulatory network in IA.

Deep venous drainage variant rate and degree may be higher in patients with perimesencephalic than in non-perimesencephalic angiogram-negative subarachnoid hemorrhage.

OBJECTIVES: The basal vein of Rosenthal (BVR) variant is a potential origin of bleeding in angiogram-negative subarachnoid hemorrhage (AN-SAH). We compared the rate and degree of BVR variants in patients with perimesencephalic AN-SAH (PAN-SAH) and non-perimesencephalic AN-SAH (NPAN-SAH). METHODS: We retrospectively reviewed the records of AN-SAH patients admitted to our hospital between 2013 and 2018. The associations between variables (baseline characteristics, clinical and radiological data, and outcome) with bleeding patterns and degree of BVR variants were analyzed. Additionally, potential predictors of positive findings on repeated digital-subtracted angiogram (DSA), rebleeding, delayed cerebral infarction (DCI), and poor outcome were further studied. RESULTS: A total of 273 patients with AN-SAH were included. The incidence rate and degree of BVR variants were significantly higher in PAN-SAH patients compared with those in NPAN-SAH patients (p < 0.001). Patients with normal bilateral BVRs are more likely to have a severe prognosis and diffused blood distribution (p < 0.05). We found an increased rate of positive findings on repeated DSA, DCI, rebleeding, and poor outcome at 3 months and 1 year after discharge (all p < 0.05) in patients with bilateral normal BVRs. Bilateral normal BVRs were considered a risk factor (predictor) of positive findings on repeated DSA, rebleeding, and poor outcome (all p < 0.05). CONCLUSIONS: PAN-SAH patients have a higher rate and degree of BVR variants compared with patients with NPAN-SAH. Those AN-SAH patients with bilateral normal BVRs are more likely to be of arterial origin and are at risk of suffering from rebleeding and a poor outcome. KEY POINTS: • Patients with PAN-SAH have a higher rate and degree of BVR variants compared with patients with NPAN-SAH, which suggested that AN-SAH patients with normal BVRs are more likely to originate from arterial bleeding. • AN-SAH patients with normal BVRs are more likely to have positive findings on repeated DSA examinations, as well as an increased incidence of rebleeding and poor outcome, which may assist and guide neurologists in selecting treatment.

Association between maternal vitamin D status with pregnancy outcomes and offspring growth in a population of Wuxi, China.

BACKGROUND AND OBJECTIVES: The role of maternal vitamin D in infantile growth remains unclear. METHODS AND STUDY DESIGN: Serum 25-hydroxyvitamin D [25(OH)D] concentrations were examined for pregnancies who visited the Affiliated Wuxi Maternity and Child Health Care Hospital of Nanjing Medical University from January 2016 to December 2017. Anthropometric measurements of corresponding offspring were performed from birth to 2 to 3 years old. Infantile body mass index (BMI) was transformed into age-, sex- and height- normalized z scores, and Latent Class Growth Mixture (LCGM) model was used to identify trajectories of BMI-Z. RESULTS: Among the 329 included pregnancy women, 109 (33.13 %), 190 (57.75%) and 30 (9.12%) were defined as vitamin D deficiency [25(OH)D <30 nmol/L], insufficiency [30 nmol/L≤25(OH)D<50 nmol/L] and sufficiency [25(OH)D ≥50 nmol/L], respectively. When compared with vitamin D sufficiency, maternal vitamin D deficiency was not associated with preterm birth [odds ratio (OR)=2.69, 95% confidence interval (95% CI)=0.57-12.80], small for gestation age (OR=0.99, 95% CI=0.29-3.46), and low birth weight (OR=1.69, 95% CI=0.34-8.51). Similarly, no significant relationships were found between maternal vitamin D concentrations and anthropometric indices (such as weight, length, BMI) during 0 to 3 years old. Furthermore, LCGM model identified two patterns of offspring growth: stable moderate BMI-Z and early transient BMI-Z groups. Maternal vitamin D levels were higher in the former group than the latter (p=0.037); however, maternal vitamin D status appeared to be unrelated with offspring BMI-Z trajectories in multivariable logistic regression models. CONCLUSIONS: Maternal vitamin D deficiency may not be related to adverse pregnancy outcomes as well as offspring growth.

The effectiveness of lumbar cerebrospinal fluid drainage in aneurysmal subarachnoid hemorrhage with different bleeding amounts.

Continuous lumbar drainage (LD) of cerebrospinal fluid can reduce the risk of aneurysmal subarachnoid hemorrhage (aSAH)-related complications. We evaluated the effectiveness of LD in aSAH patients with aneurysmal clipping and the relative benefits of different bleeding amounts. We retrospectively reviewed all consecutive aSAH patients who underwent aneurysm clipping in our hospital between January 1, 2014 and December 31, 2014. Outcomes and incidence of post-operative complications were compared between the LD group and the non-LD group in all patients and further analyzed in patients with the low modified Fisher Scale (mFS) (0-2) and high mFS (3-4). In 193 aSAH patients who underwent clipping, LD reduced the risk of hydrocephalus and improved the Glasgow Outcome Scale (GOS) score at discharge and at 3 months of follow-up. In the higher mFS group, patients who received LD had significantly lower risk of cerebral vasospasm, delayed cerebral infarction, and hydrocephalus; the GOS score was significantly higher in the LD group at discharge and at 3 months of follow-up. However, LD showed no benefits in terms of post-operative complications and outcome in patients with low mFS. LD for aneurysm clipping surgery after aSAH can reduce the risk of post-operative complications and improve the clinical outcome in patients with mFS grades 3 and 4. It should be considered as an adjunctive but dispensable treatment for aneurysm clipping in aSAH patient with low mFS.

Global Quantitative Proteomics Studies Revealed Tissue-Preferential Expression and Phosphorylation of Regulatory Proteins in Arabidopsis.

Organogenesis in plants occurs across all stages of the life cycle. Although previous studies have identified many genes as important for either vegetative or reproductive development at the RNA level, global information on translational and post-translational levels remains limited. In this study, six Arabidopsis stages/organs were analyzed using quantitative proteomics and phosphoproteomics, identifying 2187 non-redundant proteins and evidence for 1194 phosphoproteins. Compared to the expression observed in cauline leaves, the expression of 1445, 1644, and 1377 proteins showed greater than 1.5-fold alterations in stage 1-9 flowers, stage 10-12 flowers, and open flowers, respectively. Among these, 294 phosphoproteins with 472 phosphorylation sites were newly uncovered, including 275 phosphoproteins showing differential expression patterns, providing molecular markers and possible candidates for functional studies. Proteins encoded by genes preferentially expressed in anther (15), meiocyte (4), or pollen (15) were enriched in reproductive organs, and mutants of two anther-preferentially expressed proteins, acos5 and mee48, showed obviously reduced male fertility with abnormally organized pollen exine. In addition, more phosphorylated proteins were identified in reproductive stages (1149) than in the vegetative organs (995). The floral organ-preferential phosphorylation of GRP17, CDC2/CDKA.1, and ATSK11 was confirmed with western blot analysis. Moreover, phosphorylation levels of CDPK6 and MAPK6 and their interacting proteins were elevated in reproductive tissues. Overall, our study yielded extensive data on protein expression and phosphorylation at six stages/organs and provides an important resource for future studies investigating the regulatory mechanisms governing plant development.

Temperature Rise Associated with Adiabatic Shear Band: Causality Clarified.

One of the most important issues related to adiabatic shear failure is the correlation among temperature elevation, adiabatic shear band (ASB) formation and the loss of load capacity of the material. Our experimental results show direct evidence that ASB forms several microseconds after stress collapse and temperature rise reaches its maximum about 30 µs after ASB formation. This observation indicates that temperature rise cannot be the cause of ASB. Rather, it might be the result of adiabatic shear localization. As such, the traditional well-accepted thermal-softening mechanism of ASB needs to be reconsidered.

AdipoRon Protects Against Secondary Brain Injury After Intracerebral Hemorrhage via Alleviating Mitochondrial Dysfunction: Possible Involvement of AdipoR1-AMPK-PGC1α Pathway.

Intracerebral hemorrhage (ICH) is a stroke subtype that is associated with high mortality and disability rate. Mitochondria plays a crucial role in neuronal survival after ICH. This study first showed that activation of adiponectin receptor 1 (AdipoR1) by AdipoRon could attenuate mitochondrial dysfunction after ICH. In vivo, experimental ICH model was established by autologous blood injection in mice. AdipoRon was injected intraperitoneally (50 mg/kg). Immunofluorescence staining were performed to explicit the location of AdipoR1, AMP-activated protein kinase (AMPK) and peroxisome proliferator-activated receptor-γ coactivator-1a (PGC1α). The PI staining was used to quantify neuronal survival. The expression of AdipoR1 and its downstream signaling molecules were detected by Western blotting. In vitro, 10 µM oxyhemoglobin (OxyHb) was used to induce the neuronal injury in SH-SY5Y cells. Annexin V-FITC/PI staining was used to detect the neuronal apoptosis and necrosis. Mitochondrial membrane potential (Δψm) was measured by a JC-1 kit and mitochondrial mass was quantified by mitochondrial fluorescent probe. In vivo, PI staining showed that the administration of AdipoRon could reduce neuronal death at 72 h after ICH in mice. AdipoRon treatment enhanced ATP levels and reduced ROS levels in perihematoma tissues, and increased the protein expression of AdipoR1, P-AMPK, PGC1α, NRF1 and TFAM. In vitro, the JC-1 staining and Mito-tracker™ Green showed that AdipoRon significantly alleviated OxyHb-induced collapse of Δψm and enhanced mitochondrial mass. Moreover, flow cytometry analysis indicated that the neurons treated with AdipoRon showed low necrotic and apoptotic rate. AdipoRon alleviates mitochondrial dysfunction after intracerebral hemorrhage via the AdipoR1-AMPK-PGC1α pathway.

Reversible Lysine Derivatization Enabling Improved Arg-C Digestion, a Highly Specific Arg-C Digestion Using Trypsin.

The bottom-up proteomics approach has become an important strategy in diverse areas of biological research, and the enzymatic digestion is essential for this technology. Endopeptidase Arg-C catalyzing the hydrolytic cleavage of peptide bonds C-terminal to arginine could be an important protease in bottom-up proteomics. However, it has been seldom applied due to its low specificity and high cost. In this report, the reversible amine derivatization method (citraconylation and decitraconylation) was introduced and optimized toward a real Arg-C digestion using trypsin. Combination of the reversible derivatization and trypsin digestion (termed iArg-C digestion for improved Arg-C digestion) resulted in 64.2% more peptide identification (11 925 ± 199 vs 7262 ± 59) and significantly higher cleavage specificity (95.6% vs 73.6%) than the conventional Arg-C digestion. Comparison of iArg-C digestion with the widely used trypsin and Lys-C digestion revealed that iArg-C performed slightly better than Lys-C although not comparable to trypsin. Therefore, the well-established iArg-C digestion method is a promising approach for proteomics studies and could be used as the prior alternative digestion method to trypsin digestion in order to achieve higher proteome coverage. Data are available via ProteomeXchange with identifier PXD007994.

Cytological and Transcriptomic Analyses Reveal Important Roles of CLE19 in Pollen Exine Formation.

The CLAVATA3/ESR-RELATED (CLE) peptide signals are required for cell-cell communication in several plant growth and developmental processes. However, little is known regarding the possible functions of the CLEs in the anther. Here, we show that a T-DNA insertional mutant, and dominant-negative (DN) and overexpression (OX) transgenic plants of the CLE19 gene, exhibited significantly reduced anther size and pollen grain number and abnormal pollen wall formation in Arabidopsis (Arabidopsis thaliana). Interestingly, the DN-CLE19 pollen grains showed a more extensively covered surface, but CLE19-OX pollen exine exhibited clearly missing connections in the network and lacked separation between areas that normally form the lacunae. With a combination of cell biological, genetic, and transcriptomic analyses on cle19, DN-CLE19, and CLE19-OX plants, we demonstrated that CLE19-OX plants produced highly vacuolated and swollen aborted microspores (ams)-like tapetal cells, lacked lipidic tapetosomes and elaioplasts, and had abnormal pollen primexine without obvious accumulation of sporopollenin precursors. Moreover, CLE19 is important for the normal expression of more than 1,000 genes, including the transcription factor gene AMS, 280 AMS-downstream genes, and other genes involved in pollen coat and pollen exine formation, lipid metabolism, pollen germination, and hormone metabolism. In addition, the DN-CLE19(+/+) ams(-/-) plants exhibited the ams anther phenotype and ams(+/-) partially suppressed the DN-CLE19 transgene-induced pollen exine defects. These findings demonstrate that the proper amount of CLE19 signal is essential for the normal expression of AMS and its downstream gene networks in the regulation of anther development and pollen exine formation.

An optimized guanidination method for large-scale proteomic studies.

As an ε-amine specific derivatization method, guanidination is widely used in proteomic studies for mainly two reasons: the significant improvement in ionization efficiency and the selective protection of ε-amine. Herein, we employed a systematic comparison of two widely used guanidination approaches and revealed the advantages and disadvantages of each method. The sodium buffer based approach resulted in an unexpected side modification, +57 Da, which is reported for the first time; whereas the ammonium buffer based approach resulted in relatively lower yield. We carried out an optimization study by testing different buffer compositions, pH, temperatures and reaction times, and consequently discovered the optimized guanidination condition. Furthermore, we decoded the +57 Da side product as the addition of C2 H3 NO and proposed a possible mechanism of the side reaction. Importantly, our study demonstrated that mass spectrometry is a powerful tool in discovering minor side reactions which are often impossible by other techniques, and hence suggested that chemical derivatization methods should be investigated more carefully prior to extensive applications in proteomics field.

Abundant protein phosphorylation potentially regulates Arabidopsis anther development.

As the male reproductive organ of flowering plants, the stamen consists of the anther and filament. Previous studies on stamen development mainly focused on single gene functions by genetic methods or gene expression changes using comparative transcriptomic approaches, especially in model plants such as Arabidopsis thaliana However, studies on Arabidopsis anther protein expression and post-translational modifications are still lacking. Here we report proteomic and phosphoproteomic studies on developing Arabidopsis anthers at stages 4-7 and 8-12. We identified 3908 high-confidence phosphorylation sites corresponding to 1637 phosphoproteins. Among the 1637 phosphoproteins, 493 were newly identified, with 952 phosphorylation sites. Phosphopeptide enrichment prior to LC-MS analysis facilitated the identification of low-abundance proteins and regulatory proteins, thereby increasing the coverage of proteomic analysis, and facilitated the analysis of more regulatory proteins. Thirty-nine serine and six threonine phosphorylation motifs were uncovered from the anther phosphoproteome and further analysis supports that phosphorylation of casein kinase II, mitogen-activated protein kinases, and 14-3-3 proteins is a key regulatory mechanism in anther development. Phosphorylated residues were preferentially located in variable protein regions among family members, but they were they were conserved across angiosperms in general. Moreover, phosphorylation might reduce activity of reactive oxygen species scavenging enzymes and hamper brassinosteroid signaling in early anther development. Most of the novel phosphoproteins showed tissue-specific expression in the anther according to previous microarray data. This study provides a community resource with information on the abundance and phosphorylation status of thousands of proteins in developing anthers, contributing to understanding post-translational regulatory mechanisms during anther development.

Systematic Optimization of C-Terminal Amine-Based Isotope Labeling of Substrates Approach for Deep Screening of C-Terminome.

It is well-known that protein C-termini play important roles in various biological processes, and thus the precise characterization of C-termini is essential for fully elucidating protein structures and understanding protein functions. Although many efforts have been made in the field during the latest 2 decades, the progress is still far behind its counterpart, N-termini, and it necessitates more novel or optimized methods. Herein, we report an optimized C-termini identification approach based on the C-terminal amine-based isotope labeling of substrates (C-TAILS) method. We optimized the amidation reaction conditions to achieve higher yield of fully amidated product. We evaluated different carboxyl and amine blocking reagents and found the superior performance of Ac-NHS and ethanolamine. Replacement of dimethylation with acetylation for Lys blocking resulted in the identification of 232 C-terminal peptides in an Escherichia coli sample, about 42% higher than the conventional C-TAILS. A systematic data analysis revealed that the optimized method is unbiased to the number of lysine in peptides, more reproducible and with higher MASCOT scores. Moreover, the introduction of the Single-Charge Ion Inclusion (SCII) method to alleviate the charge deficiency of small peptides allowed an additional 26% increase in identification number. With the optimized method, we identified 481 C-terminal peptides corresponding to 369 C-termini in E. coli in a triplicate experiments using 80 µg each. Our optimized method would benefit the deep screening of C-terminome and possibly help discover some novel C-terminal modifications. Data are available via ProteomeXchange with identifier PXD002409.

Novel subsets of peripheral immune cells associated with promoting stroke recovery in mice.

AIMS: Peripheral immune cells infiltrating into the brain trigger neuroinflammation after an ischemic stroke. Partial immune cells reprogram their function for neural repair. Which immune cells promote ischemic brain recovery needs further identification. METHODS: We performed single-cell transcriptomic profiling of CD45high immune cells isolated from the ischemic hemisphere at subacute (5 days) and chronic (14 days) stages after ischemic stroke. RESULTS: A subset of phagocytic macrophages was associated with neuron projection regeneration and tissue remodeling. We also identified a unique type of T cells with highly expressed macrophage markers, including C1q, Apoe, Hexb, and Fcer1g, which showed high abilities in tissue remodeling, myelination regulation, wound healing, and anti-neuroinflammation. Moreover, natural killer cells decreased cytotoxicity and increased energy and metabolic function in the chronic stage after ischemic stroke. Two subgroups of neutrophils upregulated CCL signals to recruit peripheral immune cells and released CXCL2 to keep self-recruiting at the chronic stage. CONCLUSIONS: We identified subsets of peripheral immune cells that may provide potential therapeutic targets for promoting poststroke recovery.

Genome-wide association studies of peduncle length in wheat under rain-fed and irrigating field conditions.

Drought is one of the most destructive environmental factors limiting wheat production and food security globally. Peduncle length (PLE) is an important morphological trait to determine plant architecture, photosynthate transport, and yield formation, which is also considered a useful index for drought tolerance in wheat. However, the genetic basis of wheat PLE is not well studied at present. Here, a large-scale genome-wide association study (GWAS) of PLE was performed using a panel of 282 wheat accessions with the Wheat 660K SNP array genotyping under rain-fed and irrigating field conditions. Totally, 1,301 significant marker-trait associations (MTAs) were identified using the threshold of p-value < 4.16 × 10-4, five of which were high-confidence. Furthermore, combining GWAS intervals, previously reported QTLs, expression levels, homologous genes, and selected sweep analysis, a total of 5 candidate genes were detected to associate with drought stress. Moreover, the expression levels of TraesCS2A02G082100 were significantly up-regulated under drought conditions and co-localized in the selected sweep region, suggesting it is a drought-responsive gene. Our results shed light on the genetic basis underlying wheat drought tolerance, which accelerates the marker-assistant selection and genetic improvement through genomic breeding in wheat.

A novel phenotype of B cells associated with enhanced phagocytic capability and chemotactic function after ischemic stroke.

Accumulating evidence has demonstrated the involvement of B cells in neuroinflammation and neuroregeneration. However, the role of B cells in ischemic stroke remains unclear. In this study, we identified a novel phenotype of macrophage-like B cells in brain-infiltrating immune cells expressing a high level of CD45. Macrophage-like B cells characterized by co-expression of B-cell and macrophage markers, showed stronger phagocytic and chemotactic functions compared with other B cells and showed upregulated expression of phagocytosis-related genes. Gene Ontology analysis found that the expression of genes associated with phagocytosis, including phagosome- and lysosome-related genes, was upregulated in macrophage-like B cells. The phagocytic activity of macrophage-like B cells was verified by immunostaining and three-dimensional reconstruction, in which TREM2-labeled macrophage-like B cells enwrapped and internalized myelin debris after cerebral ischemia. Cell-cell interaction analysis revealed that macrophage-like B cells released multiple chemokines to recruit peripheral immune cells mainly via CCL pathways. Single-cell RNA sequencing showed that the transdifferentiation to macrophage-like B cells may be induced by specific upregulation of the transcription factor CEBP family to the myeloid lineage and/or by downregulation of the transcription factor Pax5 to the lymphoid lineage. Furthermore, this distinct B cell phenotype was detected in brain tissues from mice or patients with traumatic brain injury, Alzheimer's disease, and glioblastoma. Overall, these results provide a new perspective on the phagocytic capability and chemotactic function of B cells in the ischemic brain. These cells may serve as an immunotherapeutic target for regulating the immune response of ischemic stroke.

Dissecting human placental cells heterogeneity in preeclampsia and gestational diabetes using single-cell sequencing.

Preeclampsia (PE) and gestational diabetes mellitus (GDM) are pregnancy-specific complications, which affect maternal health and fetal outcomes. Currently, clinical and pathological studies have shown that placenta homeostasis is affected by these two maternal diseases. In this study, we aimed to gain insight into the heterogeneous changes in cell types in placental tissue-isolated from cesarean section by single-cell sequencing, including those patients diagnosed with PE (n = 5), GDM (n = 5) and healthy control (n = 5). A total of 96,048 cells (PE: 31,672; GDM: 25,294; control: 39,082) were identified in six cell types, dominated by trophoblast cells and immune cells. In addition, trophoblast cells were divided into four subtypes, including cytotrophoblast cells (CTBs), villous cytotrophoblasts (VCTs), syncytiotrophoblast (STB), and extravillous trophoblasts (EVTs). Immune cells are divided into lymphocytes and macrophages, of which macrophages have 3 subtypes (decidual macrophages, Hofbauer cells and macrophages), and lymphocytes have 4 subtypes (BloodNK, T cells, plasma cells, and decidual natural killer cells). Meanwhile, we also proved the orderly differentiation sequence of CTB into VCT, then STB and EVT. By pair-wise analysis of the expression and enrichment of differentially expressed genes in trophoblast cells between PE, GDM and control, it was found that these cells were involved in immune, nutrient transfer, hormone and oxidative stress pathways. In addition, T cells and macrophages play an immune defense role in both PE and GDM. The proportion of CTB and EVT cells in placental tissue was confirmed by flow cytometry. Taken together, our results suggested that the human placenta is a dynamic heterogenous organ dominated by trophoblast and immune cells, which perform their respective roles and interact with other cells in the environment to maintain normal placental function.