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Isolation of Group 2 Innate Lymphoid Cells from Mouse Nasal Mucosa to Detect the Expression of CD226

With abundant research on group 2 innate lymphoid cells (ILC2s) published over the years, ILC2s are widely known to be implicated in regulating various ...

Fourth Military Medical University

China is experiencing significant public health challenges related to social and demographic transitions and lifestyle transformations following unprecedented economic reforms four decades ago. Of particular public health concern is the fourfold increase in overweight and obesity rates in the nation's youth population, coupled with the low prevalence of adolescents meeting recommended levels of physical activity. Improving the overall health of China's more than 170 million children and adolescents has become a national priority. However, advancing nationwide health initiatives and physical activity promotion in this population has been hampered by the lack of a population-specific and culturally relevant consensus on recommendations for achieving these ends. To address this deficiency and inform policies to achieve  goals, a panel of Chinese experts, complemented by international professionals, developed this consensus statement. The consensus was achieved through an iterative process that began with a literature search from electronic databases; in-depth reviews, conducted by a steering committee, of the resulting articles; and panel group evaluations and discussions in the form of email correspondence, conference calls and written communications. Ultimately, the panel agreed on 10 major themes with strong scientific evidence that, in children and adolescents aged 6-17, participating in moderate to vigorous physical activities led to multiple positive health outcomes. Our consensus statement also (1) highlights major challenges in promoting physical activity, (2) identifies future research that addresses current knowledge gaps, and (3) provides recommendations for teachers, education experts, parents and policymakers for promoting physical activity among Chinese school-aged children and adolescents. This consensus statement aligns with international efforts to develop global physical activity guidelines to promote physical activity and health and prevent lifestyle-related diseases in children and adolescents. More importantly, it provides a foundation for developing culturally appropriate and effective physical activity interventions, health promotion strategies and policy initiatives to improve the health of Chinese children and adolescents.

Physical activity and health in Chinese children and adolescents: expert consensus statement (2020).

The present study aims to delineate the working mechanism of prediction in sentence comprehension, by disentangling the influence of the facilitated general memory retrieval from the coexistent influence of the predicted language-specific semantic and/or syntactic information for the first time. The results support that prediction might influence the downstream cognitive processing in two aspects: (1) the pre-activated information facilitates the retrieval of a matched input in memory and, (2) the pre-activated information interacts with higher-level semantic/syntactic processing. More importantly, the present findings suggest that these two types of influences seem to occur at different stages of sentence comprehension: the facilitated memory retrieval of the input modulates N400 amplitude and the latency of post-N400 late central-parietal positivity/P600, while the predicted semantic/syntactic information and/or their interactions modulate the amplitude of the late positivity. The present findings would be helpful for interpreting the underlying mechanism of observed effects in prediction studies.

Delving Into the Working Mechanism of Prediction in Sentence Comprehension: An ERP Study.

Unlike in English, the role of phonology in word recognition in Chinese is unclear. In this event-related potential experiment, we investigated the role of phonology in reading both high- and low-frequency two-character compound Chinese words. Participants executed semantic and homophone judgment tasks of the same precede-target pairs. Each pair of either high- or low-frequency words were either unrelated (control condition) or related semantically or phonologically (homophones). The induced P200 component was greater for low- than for high-frequency word-pairs both in semantic and phonological tasks. Homophones in the semantic judgment task and semantically-related words in the phonology task both elicited a smaller N400 than the control condition, word frequency-independently. However, for low-frequency words in the phonological judgment task, it was found that the semantically related pairs released a significantly larger P200 than the control condition. Thus, the semantic activation of both high- and low-frequency words may be no later than phonological activation.

An ERP Study on the Role of Phonological Processing in Reading Two-Character Compound Chinese Words of High and Low Frequency.

Osteosarcoma (OS) is a highly malignant and aggressive bone tumor. This study was performed to explore the mechanisms of HuR (human antigen R) in the progression of OS.

Silencing of HuR Inhibits Osteosarcoma Cell Epithelial-Mesenchymal Transition  AGO2 in Association With Long Non-Coding RNA XIST.

Platelet-rich plasma (PRP) is a platelet-based application used to treat osteoarthritis (OA) clinically. The co-stimulatory molecule CD226 is expressed in T cells, NK cells, and also platelets. However, exact effects of CD226 on platelets and whether its expression level influences PRP efficacy are largely unknown. Here, CD226PF4-Cre mice were obtained from mating CD226 mice with PF4-Cre mice. Blood samples and washed platelets were collected from the mice eyeballs to undergo routine blood tests and transmission electron microscopy. Differentially expressed proteins were detected by iTRAQ-based proteomics analysis. Animal OA models were established through surgical destabilization of the medial meniscus (DMM) for C57BL/6 wildtype mice, followed by PRP injection to evaluate the effects of platelet CD226 on PRP efficacy. The results showed that deletion of platelet CD226 increased the number of megakaryocytes (MKs) in bone marrow (BM) but reduced MKs in spleen, combined with significantly decreased platelet amounts, α-granule secretion, and reduced immature platelets; indicating that absence of platelet CD226 may disrupt MK/platelet homeostasis and arrested platelet release from MKs. Sequencing analysis showed abnormal ribosomal functions and much downregulated proteins in the absence of platelet CD226. Autophagy-related proteins were also reduced in the CD226-absent MKs/platelets. Moreover, deletion of platelet CD226 diminished the protective effects of PRP on DMM-induced cartilage lesions in mice, and PDGF restored it. Therefore, deficiency of platelet CD226 inhibited platelet maturation, secretion, and normal ribosomal functions, which may lead to depressed PRP efficacy on OA, suggesting that CD226 is required to regulate platelet growth, functions, and its application.

CD226 Is Required to Maintain Megakaryocytes/Platelets Homeostasis in the Treatment of Knee Osteoarthritis With Platelet-Rich Plasma in Mice.

Hemorrhagic shock (HS) is common in clinical emergencies, leading to millions of deaths each year globally. CD226 is a costimulatory adhesion molecule expressed on both immune cells and endothelial cells (ECs) to regulate their metabolic activity and function. As endothelial dysfunction occurs after HS, the roles CD226 plays in vascular EC metabolism were investigated. CD226 Tek mice were adopted to achieve vascular EC-specific knockout of CD226, and subjected to HS modelling. Serum levels of crucial intermediate metabolites were evaluated through liquid chromatography-mass spectrometry analysis. Human umbilical vein ECs (HUVECs) were used to study the effects of CD226 under hypoxia in vitro. Seahorse analysis evaluated the cellular glycolysis and mitochondria bioenergetics. Results showed that CD226 deficiency in vascular ECs alleviated HS-induced intestinal damage and inflammatory response in mice. Animal studies indicated an improved energy metabolism when CD226 was knocked out in ECs after HS, as evidenced by enhanced glutamine-glutamate metabolism and decreased lactic acid levels. Glut-1 was upregulated in mouse vascular ECs after HS and HUVECs under hypoxia, combined with decreased CD226. Moreover, HUVECs with CD226 knockdown exhibited relieved mitochondrial damage and early apoptosis under hypoxia, whereas CD226 overexpression showed opposite effects. Seahorse analysis showed that downregulated CD226 significantly increased mitochondrial ATP production and glucose uptake in HUVECs under hypoxia. Additionally, Erk/PHD2 signaling-mediated HIF-1α/Glut-1 and HIF-2α/ASCT2 pathways were involved in CD226 regulation on HUVEC glutaminolysis after hypoxia. Hence, CD226 deficiency promotes bypass energy supply to vascular ECs under ischemic or hypoxic stress, to ameliorate the stress-mediated metabolic disturbance.

CD226 deficiency promotes glutaminolysis and alleviates mitochondria damage in vascular endothelial cells under hemorrhagic shock.

Growing evidence has demonstrated that stress triggers gastrointestinal (GI) disorders. This study aimed to investigate how the acute cold water-immersion restraint (CWIR) stress affects intestinal injury and gut microbiota (GM) distribution. Male C57BL/6 mice were used to establish a CWIR animal model. Hematoxylin-eosin and periodic acid-Schiff staining were performed to assess intestinal histopathological changes. Reverse transcription quantitative polymerase chain reaction (RT-qPCR) analysis and immunofluorescence staining were used to evaluate the expression of inflammatory cytokines and immune cell infiltration in the intestinal tissues. The gut permeability and intestinal occludin protein expression were determined through fluorescein isothiocyanate-dextran detection and western blot, respectively. GM profiles were analyzed  high-throughput sequencing of the fecal bacterial 16S rRNA genes. Results showed that CWIR induced more severe intestinal mucosal injury compared to the control, leading to a significant increase in tumor necrosis factor-α expression, but no infiltration of neutrophil and T cells. CWIR also resulted in GI disruption and increased the permeability of the intestinal mucosa. GM profiles showed that CWIR reduced GM diversity of mice compared with the control group. Specifically, aerobic and gram-negative bacteria significantly increased after CWIR, which was associated with the severity of gut injury under stress. Therefore, acute CWIR leads to severe intestinal damage with inflammation and disrupts the GM homeostasis, contributing to decreased GM diversity. Our findings provide the theoretical basis for the further treatment of intestinal disorders induced by CWIR.

Acute Cold Water-Immersion Restraint Stress Induces Intestinal Injury and Reduces the Diversity of Gut Microbiota in Mice.

Obesity is associated with chronic low-grade inflammation, contributing to an increasing prevalence of chronic metabolic diseases, such as insulin resistance, non-alcoholic fatty liver disease (NALFD), and steatohepatitis. Macrophages are the predominant immune cells in adipose tissues. Adipose tissue macrophages (ATMs) would switch to pro-inflammatory M1 state during obesity, causing local and systemic inflammation. However, the regulatory mechanism of ATMs has not yet been well described within this process. Using a high-fat diet (HFD)-induced mouse obesity model, we found that the costimulatory molecule CD226 was highly expressed on ATMs and knockout (KO) of CD226 alleviated obesity caused by HFD. Loss of CD226 reduced the accumulation of ATMs and hindered macrophage M1 polarization, with lower serum proinflammatory cytokine levels. Furthermore, deficiency of CD226 on ATMs decreased the phosphorylation levels of VAV1, AKT, and FOXO1 and thereby upregulated PPAR-γ. Further administration of PPAR-γ inhibitor restored M1 phenotype in CD226KO ATMs. In summary, loss of CD226 alleviates the HFD-induced obesity and systemic inflammation through inhibition of the accumulation and M1 polarization of ATMs in which PPAR-γ-dependent signaling pathway is involved, suggesting that CD226 may be identified as a potential molecular target for the clinical treatment of obesity.

CD226 knockout alleviates high-fat diet induced obesity by suppressing proinflammatory macrophage phenotype.

Interleukin-6 (IL-6) in mucosal immune cells is involved in post-injury intestinal regeneration, inflammation responses, and gastric homeostasis. However, the interaction between IL-6 and the dynamic balance of gut microbiota (GM) remains unexplored. Intestinal pathology was assessed by hematoxylin and eosin and periodic acid-Schiff staining in wild-type (WT) and IL-6 gene knockout (KO) C57BL/6J mice. GM profiles were established via high-throughput sequencing of the fecal bacterial 16S rRNA gene. Intestinal α- and β-defensins were measured by quantitative real-time PCR; further, flow cytometry was performed to analyze isolated intraepithelial lymphocytes (IELs). Compared with the WT, IL-6 KO did not obviously change gut structures, but significantly reduced GM diversity, resulting in reduced metabolic pathways with decreased gram-positive but elevated gram-negative bacteria. More taxa alterations included differences at the phyla (e.g., increased Verrucomicrobia and decreased Firmicutes) and genera (e.g., increased Akkermansia and decreased Lactobacillus) levels. Absence of IL-6 also significantly increased intestinal expression of defensins α3 and α4 (Defa3 and Defa4) and the percentage of natural TCRγδ IELs, providing a molecular basis for triggering mucosal immune response. Therefore, IL-6 loss remodels GM composition and alters IEL maintenance, identifying IL-6 as a crucial cytokine for GM dysbiosis and mucosal immunity.

Article	Year
Physical activity and health in Chinese children and adolescents: expert consensus statement (2020). British journal of sports medicine\| PubMed ID: 32471813	2020
Delving Into the Working Mechanism of Prediction in Sentence Comprehension: An ERP Study. Frontiers in psychology\| PubMed ID: 33679524	2021
An ERP Study on the Role of Phonological Processing in Reading Two-Character Compound Chinese Words of High and Low Frequency. Frontiers in psychology\| PubMed ID: 33716906	2021
Silencing of HuR Inhibits Osteosarcoma Cell Epithelial-Mesenchymal Transition AGO2 in Association With Long Non-Coding RNA XIST. Frontiers in oncology\| PubMed ID: 33816232	2021
CD226 Is Required to Maintain Megakaryocytes/Platelets Homeostasis in the Treatment of Knee Osteoarthritis With Platelet-Rich Plasma in Mice. Frontiers in pharmacology\| PubMed ID: 34526904	2021
CD226 deficiency promotes glutaminolysis and alleviates mitochondria damage in vascular endothelial cells under hemorrhagic shock. FASEB journal : official publication of the Federation of American Societies for Experimental Biology\| PubMed ID: 34669985	2021
Acute Cold Water-Immersion Restraint Stress Induces Intestinal Injury and Reduces the Diversity of Gut Microbiota in Mice. Frontiers in cellular and infection microbiology\| PubMed ID: 34722327	2021
CD226 knockout alleviates high-fat diet induced obesity by suppressing proinflammatory macrophage phenotype. Journal of translational medicine\| PubMed ID: 34823548	2021
Interleukin-6 absence triggers intestinal microbiota dysbiosis and mucosal immunity in mice. Cytokine\| PubMed ID: 35276634	2022

Yuling Wang

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