A novel sponge composite of chitosan-sodium tripolyphosphate-melamine for anionic dye Orange II removal.

Since the potential carcinogenic, toxic and non-degradable dyes trigger serious environmental contamination by improper treatment, developing novel adsorbents remains a major challenge. A novel high efficiency and biopolymer-based environmental-friendly adsorbent, chitosan­sodium tripolyphosphate-melamine sponge (CTS-STPP-MS) composite, was prepared for Orange II removing with chitosan as raw material, sodium tripolyphosphate as cross-linking agent. The composite was carefully characterized by SEM, EDS, FT-IR and XPS. The influence of crosslinking conditions, dosage, pH, initial concentration, contacting time and temperature on adsorption were tested through batch adsorption experiments. CTS-STPP-MS adsorption process was exothermic, spontaneous and agreed with Sips isotherm model accompanying the maximum adsorption capacity as 948 mg∙g-1 (pH = 3). Notably, the adsorption performance was outstanding for high concentration solutions, with a removal rate of 97 % in up to 2000 mg∙L-1 OII solution (100 mg sorbent dosage, 50 mL OII solution, pH = 3, 289.15 K). In addition, the adsorption efficiency yet remained 97.85 % after 5 repeated adsorption-desorption cycles. The driving force of adsorption was attributed to electrostatic attraction and hydrogen bonds which was proved by adsorption results coupled with XPS. Owing to the excellent properties of high-effective, environmental-friendly, easy to separate and regenerable, CTS-STPP-MS composite turned out to be a promising adsorbent in contamination treatment.

GDF9His209GlnfsTer6/S428T and GDF9Q321X/S428T bi-allelic variants caused female subfertility with defective follicle enlargement.

BACKGROUND: Antral follicles consist of an oocyte cumulus complex surrounding by somatic cells, including mural granulosa cells as the inner layer and theca cells as the outsider layer. The communications between oocytes and granulosa cells have been extensively explored in in vitro studies, however, the role of oocyte-derived factor GDF9 on in vivo antral follicle development remains elusive due to lack of an appropriate animal model. Clinically, the phenotype of GDF9 variants needs to be determined. METHODS: Whole-exome sequencing (WES) was performed on two unrelated infertile women characterized by an early rise of estradiol level and defect in follicle enlargement. Besides, WES data on 1,039 women undergoing ART treatment were collected. A Gdf9Q308X/S415T mouse model was generated based on the variant found in one of the patients. RESULTS: Two probands with bi-allelic GDF9 variants (GDF9His209GlnfsTer6/S428T, GDF9Q321X/S428T) and eight GDF9S428T heterozygotes with normal ovarian response were identified. In vitro experiments confirmed that these variants caused reduction of GDF9 secretion, and/or alleviation in BMP15 binding. Gdf9Q308X/S415T mouse model was constructed, which recapitulated the phenotypes in probands with abnormal estrogen secretion and defected follicle enlargement. Further experiments in mouse model showed an earlier expression of STAR in small antral follicles and decreased proliferative capacity in large antral follicles. In addition, RNA sequencing of granulosa cells revealed the transcriptomic profiles related to defective follicle enlargement in the Gdf9Q308X/S415T group. One of the downregulated genes, P4HA2 (a collagen related gene), was found to be stimulated by GDF9 protein, which partly explained the phenotype of defective follicle enlargement. CONCLUSIONS: GDF9 bi-allelic variants contributed to the defect in antral follicle development. Oocyte itself participated in the regulation of follicle development through GDF9 paracrine effect, highlighting the essential role of oocyte-derived factors on ovarian response.

Research on the Low-Temperature Impact Toughness of a New 100-mm Ultra-Thick Offshore Steel Fabricated Using the Narrow-Gap Laser Wire Filling Welding Process.

Ultra-thick offshore steel, known for its high strength, high toughness, and corrosion resistance, is commonly used in marine platforms and ship components. However, when offshore steel is in service for an extended period under conditions of high pressure, extreme cold, and high-frequency impact loads, the weld joints are prone to fatigue failure or even fractures. Addressing these issues, this study designed a narrow-gap laser wire filling welding process and successfully welded a 100-mm new type of ultra-thick offshore steel. Using finite element simulation, EBSD testing, SEM analysis, and impact experiments, this study investigates the weld's microstructure, impact toughness, and fracture mechanisms. The research found that at -80 °C, the welded joint exhibited good impact toughness (>80 J), with the impact absorption energy on the surface of the weld being 217.7 J, similar to that of the base material (225.3 J), and the fracture mechanism was primarily a ductile fracture. The impact absorption energy in the core of the weld was 103.7 J, with the fracture mechanism mainly being a brittle fracture. The EBSD results indicated that due to the influence of the welding thermal cycle and the cooling effect of the narrow-gap process, the grains gradually coarsened from the surface of the welded plate to the core of the weld, which was the main reason for the decreased impact toughness at the joint core. This study demonstrates the feasibility of using narrow-gap laser wire filling welding for 100-mm new type ultra-thick offshore steel and provides a new approach for the joining of ultra-thick steel plates.

Fine tuning of CpG spatial distribution with DNA origami for improved cancer vaccination.

Multivalent presentation of ligands often enhances receptor activation and downstream signalling. DNA origami offers a precise nanoscale spacing of ligands, a potentially useful feature for therapeutic nanoparticles. Here we use a square-block DNA origami platform to explore the importance of the spacing of CpG oligonucleotides. CpG engages Toll-like receptors and therefore acts to activate dendritic cells. Through in vitro cell culture studies and in vivo tumour treatment models, we demonstrate that square blocks induce Th1 immune polarization when CpG is spaced at 3.5 nm. We observe that this DNA origami vaccine enhances DC activation, antigen cross-presentation, CD8 T-cell activation, Th1-polarized CD4 activation and natural-killer-cell activation. The vaccine also effectively synergizes with anti-PD-L1 for improved cancer immunotherapy in melanoma and lymphoma models and induces long-term T-cell memory. Our results suggest that DNA origami may serve as a platform for controlling adjuvant spacing and co-delivering antigens in vaccines.

Diesel soot combustion in air-NO environment: Evolution of functional groups on soot surfaces.

It is inevitable for NO to be involved in the soot combustion in diesel particulate filters (DPFs), so giving full play to the NO oxidation activity is one of the most effective means to improve the DPF regeneration performance. In this work, based on the results of programmed temperature oxidation (TPO) experiments, Fourier transfer inference spectroscopy, and X-ray photoelectron spectroscopy, the evolution of surface functional groups was seriously analyzed to explore the soot oxidation mechanism. The results revealed that with the presence of NO in the air atmosphere, the concentration of -ONO2 groups showed an increasing trend in the early oxidation stage of 0-20 % oxidation degree (OD) and then slowly decreased during 20-80 % OD, while the variations in CH functional group concentration were directly related to the concentration NO in the air atmosphere. COO functional group is easy to decompose, and NO promotes COO's generation and decomposition. The sp3/sp2 hybrid ratio is strongly correlated with CO (carbon­oxygen double bond), but the content of CO is also affected by the desorption of COO functional groups. It is worth noting that when the soot oxidation degree is at 50 %-80 % OD, CO groups are converted to CO functional groups.

Methyl donor diet attenuates intimal hyperplasia after vascular injury in rats.

Environmental factors, particularly dietary habits, play an important role in cardiovascular disease susceptibility and progression through epigenetic modification. Previous studies have shown that hyperplastic vascular intima after endarterectomy is characterized by genome-wide hypomethylation. The purpose of this study was to investigate whether methyl donor diet affects intimal hyperplasia and the possible mechanisms involved. Intimal hyperplasia was induced in SD rats by carotid artery balloon injury. From 8 d before surgery to 28 d after surgery, the animals were fed a normal diet (ND) or a methyl donor diet (MD) supplemented with folic acid, vitamin B12, choline, betaine, and zinc. Carotid artery intimal hyperplasia was observed by histology, the effect of MD on carotid protein expression was analyzed by proteomics, functional clustering, signaling pathway, and upstream-downstream relationship of differentially expressed proteins were analyzed by bioinformatics. Results showed that MD attenuated balloon injury-induced intimal hyperplasia in rat carotid arteries. Proteomic analysis showed that there were many differentially expressed proteins in the common carotid arteries of rats fed with two different diets. The differentially expressed proteins are mainly related to the composition and function of the extracellular matrix (EMC), and changes in the EMC can lead to vascular remodeling by affecting fibrosis and stiffness of the blood vessel wall. Changes in the levels of vasculotropic proteins such as S100A9, ILF3, Serpinh1, Fbln5, LOX, HSPG2, and Fmod may be the reason why MD attenuates intimal hyperplasia. Supplementation with methyl donor nutrients may be a beneficial measure to prevent pathological vascular remodeling after injury.

Effect of Proinflammatory S100A9 Protein on Migration and Proliferation of Microglial Cells.

Alzheimer's disease (AD) is a multifactorial disease affecting aging population worldwide. Neuroinflammation became a focus of research as one of the major pathologic processes relating to the disease onset and progression. Proinflammatory S100A9 is the central culprit in the amyloid-neuroinflammatory cascade implicated in AD and other neurodegenerative diseases. We studied the effect of S100A9 on microglial BV-2 cell proliferation and migration. The responses of BV-2 cells to S100A9 stimulation were monitored in real-time using live cell microscopy, transcriptome sequencing, immunofluorescence staining, western blot analysis, and ELISA. We observed that a low dose of S100A9 promotes migration and proliferation of BV-2 cells. However, acute inflammatory condition (i.e., high S100A9 doses) causes diminished cell viability; it is uncovered that S100A9 activates TLR-4 and TLR-7 signaling pathways, leading to TNF-α and IL-6 expression, which affect BV-2 cell migration and proliferation in a concentration-dependent manner. Interestingly, the effects of S100A9 are not only inhibited by TNF-α and IL-6 antibodies. The addition of amyloid-ß (Aß) 1-40 peptide resumes the capacities of BV-2 cells to the level of low S100A9 concentrations. Based on these results, we conclude that in contrast to the beneficial effects of low S100A9 dose, high S100A9 concentration leads to impaired mobility and proliferation of immune cells, reflecting neurotoxicity at acute inflammatory conditions. However, the formation of Aß plaques may be a natural mechanism that rescues cells from the proinflammatory and cytotoxic effects of S100A9, especially considering that inflammation is one of the primary causes of AD.

Selenium regulates T cell differentiation in experimental autoimmune thyroiditis in mice.

Selenium (Se) is an essential trace element that plays an important role in thyroid physiology. Se supplementation can reduce levels of autoimmune thyroid antibodies, which may be beneficial in Hashimoto's thyroiditis (HT). However, the long-term benefits of Se supplementation for HT patients are controversial and there is no clear clinical evidence to support it, so further basic and clinical research is needed. The effect of Se on immune cells, especially T cells, in autoimmune thyroiditis (AIT) has not been elucidated. Here, we replicated a mouse model of experimental autoimmune thyroiditis (EAT) on a high-iodine diet and treated it with Se supplementation. At week 8 of the experiment, Se supplementation reduced the destruction of thyroid follicles and the infiltration rate of lymphocytes in EAT mice, and reversed the disturbance of peripheral blood thyroxine and thyroid autoantibody levels. Further examination revealed that Se had broad effects on T-cell subsets. Its effects include reducing the production of pro-inflammatory cytokines by Th1 cells, inhibiting the differentiation and production of cytokines by Th2 and Th17 cells, and upregulating the differentiation and production of cytokines by Treg cells. These changes help alleviate thyroid follicle damage during EAT. In conclusion, selenium supplementation has the potential to improve the prognosis of AIT by altering the subset differentiation and/or function of CD4+ T cells.

MiR-146b-5p regulates the scavenging effect of GPx-3 on peroxide in papillary thyroid cancer cells.

Background: Glutathione peroxidase (GPx) is an important antioxidant enzyme in thyroid follicular cells. Reduced levels of glutathione peroxidase 3 (GPx-3) expression in papillary thyroid cancer (PTC) are associated with poor prognosis. However, the reason for the decreased expression level of GPx-3 in PTC is unclear. Methods: The expression of GPx-3 in papillary thyroid carcinoma and adjacent normal tissue (n = 18) was detected by Western blotting. Bioinformatics was used to predict the relationship between the level of GPx-3 and gender, age, lymph node metastasis, stage, BRAFV600E mutation, and recurrence-free survival of patients. The possible upstream microRNAs of GPx-3 were analyzed by bioinformatics tools also. We verified the relationship between GPx-3 and upstream microRNA by dual luciferase reporter assay and enzyme-linked immunosorbent assay (ELISA). Results: The protein level of GPx-3 decreased in PTC, and analysis of public database datasets suggests that its decreased expression may be associated with the BRAFV600E mutation. MiR-146b-5p was significantly overexpressed in PTC. The dual luciferase reporter assay verified the effect of miR-146b-5p on 3'-UTR of GPx-3 mRNA. Knockdown of miR-146b-5p in thyroid cancer cell lines TPC-1 and BCPAP increased GPx-3 expression levels, accompanied by an increase in the conversion of glutathione (GSH) to oxidized glutathione (GSSG). Conclusions: In conclusion, the level of GPx-3 decreases in papillary thyroid carcinoma and impairs intracellular peroxide clearance, due to the inhibitory effect of miR-146b-5p. The accumulation of intracellular peroxides may contribute to the poor prognosis of thyroid cancer.

In Situ Encapsulation of Camptothecin by Self-Assembly of Poly(acrylic acid)-b-Poly(N-Isopropylacrylamide) and Chitosan for Controlled Drug Delivery.

Camptothecin (CPT) has been shown to exhibit anticancer activity against several cancers. Nevertheless, CPT is very hydrophobic with poor stability, and thus its medical application is limited. Therefore, various drug carriers have been exploited for effectively delivering CPT to the targeted cancer site. In this study, a dual pH/thermo-responsive block copolymer of poly(acrylic acid-b-N-isopropylacrylamide) (PAA-b-PNP) was synthesized and applied to encapsulate CPT. At temperatures above its cloud point, the block copolymer self-assembled to form nanoparticles (NPs) and in situ encapsulate CPT, owing to their hydrophobic interaction as evidenced by fluorescence spectrometry. Chitosan (CS) was further applied on the surface through the formation of a polyelectrolyte complex with PAA for improving biocompatibility. The average particle size and zeta potential of the developed PAA-b-PNP/CPT/CS NPs in a buffer solution were 168 nm and -30.6 mV, respectively. These NPs were still stable at least for 1 month. The PAA-b-PNP/CS NPs exhibited good biocompatibility toward NIH 3T3 cells. Moreover, they could protect the CPT at pH 2.0 with a very slow-release rate. At pH 6.0, these NPs could be internalized by Caco-2 cells, followed by intracellular release of the CPT. They became highly swollen at pH 7.4, and the released CPT was able to diffuse into the cells at higher intensity. Among several cancer cell lines, the highest cytotoxicity was observed for H460 cells. As a result, these environmentally-responsive NPs have the potential to be applied in oral administration.

GSK-3484862 targets DNMT1 for degradation in cells.

Maintenance of genomic methylation patterns at DNA replication forks by DNMT1 is the key to faithful mitotic inheritance. DNMT1 is often overexpressed in cancer cells and the DNA hypomethylating agents azacytidine and decitabine are currently used in the treatment of hematologic malignancies. However, the toxicity of these cytidine analogs and their ineffectiveness in treating solid tumors have limited wider clinical use. GSK-3484862 is a newly-developed, dicyanopyridine containing, non-nucleoside DNMT1-selective inhibitor with low cellular toxicity. Here, we show that GSK-3484862 targets DNMT1 for protein degradation in both cancer cell lines and murine embryonic stem cells (mESCs). DNMT1 depletion was rapid, taking effect within hours following GSK-3484862 treatment, leading to global hypomethylation. Inhibitor-induced DNMT1 degradation was proteasome-dependent, with no discernible loss of DNMT1 mRNA. In mESCs, GSK-3484862-induced Dnmt1 degradation requires the Dnmt1 accessory factor Uhrf1 and its E3 ubiquitin ligase activity. We also show that Dnmt1 depletion and DNA hypomethylation induced by the compound are reversible after its removal. Together, these results indicate that this DNMT1-selective degrader/inhibitor will be a valuable tool for dissecting coordinated events linking DNA methylation to gene expression and identifying downstream effectors that ultimately regulate cellular response to altered DNA methylation patterns in a tissue/cell-specific manner.

Obesity and cancer: Mouse models used in studies.

There is increasing evidence that obesity is associated with the occurrence and development of malignant tumors. When studying the relationship between obesity and malignant tumors, it is very important to choose an appropriate animal model. However, BALB/c nude mice and other animals commonly used to study tumor xenograft (human-derived tumor cell lines) transplantation models are difficult to induce obesity, while C57BL/6 mice and other model animals commonly used for obesity research are not suitable for tumor xenograft transplantation. Therefore, it is difficult to replicate both obesity and malignancy in animal models at the same time. This review summarizes several experimental animal models and protocols that can simultaneously induce obesity and tumor xenografts.

Prenatal phenotype of Wolf-Hirschhorn syndrome: A case series and literature review.

OBJECTIVE: Wolf-Hirschhorn syndrome (WHS) is a congenital malformation syndrome with poor prognosis. It is associated with a heterozygous deletion of chromosome 4p16.3. Adequate knowledge of prenatal phenotypes and proper prenatal counseling are essential for intrauterine diagnosis. METHOD: We retrospectively analyzed 11 prenatal cases of WHS diagnosed using low-depth whole-genome sequencing (copy number variation sequencing) performed at our hospital from May 2017 to September 2022 and reviewed their prenatal ultrasound reports in detail. We also analyzed WHS cases (including prenatal and postnatal) with abnormal prenatal ultrasound findings in the published literature over the past 20 years. RESULTS: Among the 11 fetuses with a prenatal diagnosis of WHS in our hospital, four cases showed abnormal prenatal ultrasound findings, including shrunken kidneys, ventricular septal defect, a small stomach, fetal growth restriction (FGR), enlarged posterior fossa, and soft ultrasonic markers. Our four cases were combined with 114 published WHS cases with prenatal ultrasound abnormalities from other medical institutions. Of the 118 cases, 59.3% (70 of 118) were multiple malformations. The most frequent ultrasound features observed in all 118 cases were FGR (76.3%, 90 of 118), followed by facial anomalies (28.8%, 34 of 118), central nervous system anomalies (27.1%, 32 of 118), and soft ultrasound markers (23.7%, 28 of 118). Other less common phenotypes included cardiac anomalies (19.5%, 23 of 118), genitourinary anomalies (19.5%, 23 of 118), increased NT/NF (12.7%, 15 of 118), skeletal anomalies (11.9%, 14 of 118), a single umbilical artery (10.2%, 12 of 118), gastrointestinal anomalies (9.3%, 11 of 118), oligohydramnios (8.5%, 10 of 118), cystic hygroma (5.1%, six of 118), hydrops/pleural effusion/ascites (2.5%, three of 118), and polyhydramnios (2.5%, three of 118). CONCLUSION: This study improved our understanding of the prenatal presentation of WHS by analyzing prenatal ultrasound abnormalities. The timely identification of prenatal ultrasound abnormalities can provide accurate consultation for pregnant women, improve the prenatal detection of WHS, and enable early prenatal management and intervention of WHS.

A novel salivary effector, BtE3, is essential for whitefly performance on host plants.

The whitefly Bemisia tabaci is a piercing-sucking herbivore that reduces the yields of crops both by feeding on plants and transmitting plant viruses. Like most plant feeders, B. tabaci has evolved ways to avoid plant defence responses. For example, B. tabaci is known to secrete salivary effectors to suppress host defences. However, the nature of B. tabaci effectors is not completely understood. In this study, we used B. tabaci genomic and salivary gland transcriptomic data and an overexpression system to identify a previously unknown B. tabaci salivary effector, BtE3. BtE3 is specifically expressed in the head (containing primary salivary glands) and is secreted into hosts during B. tabaci feeding. In planta overexpression of BtE3 blocked Burkholderia glumae-induced hypersensitive response (HR) in both Nicotiana benthamiana and Solanum lycopersicum. Silencing of BtE3 by plant-mediated RNAi prevented B. tabaci from continuously ingesting phloem sap, and reduced B. tabaci survival and fecundity. Moreover, overexpression of BtE3 in planta up-regulated the salicylic acid- (SA-) signalling pathway, but suppressed the downstream jasmonic acid- (JA-) mediated defences. Taken together, these results indicate that BtE3 is a B. tabaci-specific novel effector involved in B. tabaci-plant interactions. These findings increase our understanding of B. tabaci effectors and suggest novel strategies for B. tabaci pest management.

A Stand-Alone Microfluidic Chip for Long-Term Cell Culture.

Live-cell microscopy is crucial for biomedical studies and clinical tests. The technique is, however, limited to few laboratories due to its high cost and bulky size of the necessary culture equipment. In this study, we propose a portable microfluidic-cell-culture system, which is merely 15 cm×11 cm×9 cm in dimension, powered by a conventional alkali battery and costs less than USD 20. For long-term cell culture, a fresh culture medium exposed to 5% CO2 is programmed to be delivered to the culture chamber at defined time intervals. The 37 °C culture temperature is maintained by timely electrifying the ITO glass slide underneath the culture chamber. Our results demonstrate that 3T3 fibroblasts, HepG2 cells, MB-231 cells and tumor spheroids can be well-maintained for more than 48 h on top of the microscope stage and show physical characters (e.g., morphology and mobility) and growth rate on par with the commercial stage-top incubator and the widely adopted CO2 incubator. The proposed portable cell culture device is, therefore, suitable for simple live-cell studies in the lab and cell experiments in the field when samples cannot be shipped.

Impaired Cognitive Function in Patients With Autonomous Cortisol Secretion in Adrenal Incidentalomas.

CONTEXT: Glucocorticoids have potent effects on the central nervous system. However, while patients with Cushing syndrome frequently report impairments in cognitive function, studies investigating cognitive function in patients with autonomous cortisol secretion (ACS) in adrenal incidentalomas (AIs) are scarce. OBJECTIVE: The aim of the present study was to evaluate neurocognitive function in patients with ACS. METHODS: We prospectively recruited 63 patients with AI, 36 patients with nonfunctional adrenal adenoma (NFA) (46.5 ± 10.5 years), and 27 patients with ACS (48.6 ± 9.1 years); these patients underwent a battery of validated neuropsychological tests. ACS was diagnosed when serum cortisol levels after a 1-mg dexamethasone suppression test (cortisol1 mg DST) ≥ 50 nmol/L. RESULTS: Patients with ACS had higher frequency of subjective memory complaints (40.7% vs 13.9%, P < 0.05) and higher proportion of mild cognitive impairment (22.2% vs 2.8%, P < 0.05) than patients with NFA. Furthermore, patients with ACS had worse performance on working memory and the visuospatial/constructional domain than patients with NFA (all P < 0.05). Serum cortisol1 mg DST was negatively correlated with working memory and visuospatial/constructional domains (r = -0.307 and -0.306, respectively, all P < 0.05). Performance on working memory and visuospatial/constructional domains gradually deteriorated with increases in serum cortisol1 mg DST (all P values for trend < 0.05). Multivariate linear regression analysis showed that serum cortisol1 mg DST was a significant risk factor for impairment of working memory and visuospatial/constructional domains (B = -0.006 and -0.043, respectively, all P < 0.05). CONCLUSION: This study is the first to report that ACS is accompanied by impaired cognitive function. Consequently, cognitive function assessment should be incorporated into the clinical evaluation of patients with ACS. CLINICAL TRIAL REGISTRATION NUMBER: NCT05357456.

Uncovering the emergence of HSCs in the human fetal bone marrow by single-cell RNA-seq analysis.

During fetal development, human hematopoietic stem cells (HSCs) colonize the bone marrow (BM), where they self-renew and sustain hematopoiesis throughout life; however, the precise timepoint at which HSCs seed the BM is unclear. We used single-cell RNA-sequencing to map the transcriptomic landscape of human fetal BM and spleen hematopoietic stem/progenitor cells (HSPCs) and their microenvironment from 10 to 14 post-conception weeks (PCWs). We further demonstrated that functional HSCs capable of reconstituting long-term multi-lineage hematopoiesis in adult NOG mice do not emerge in the BM until 12 PCWs. In contrast, functional HSCs were not detected in the spleen by 14 PCWs. By comparing the niche-HSPC interactions between BM and spleen, we identified ligand-receptor pairs likely to be involved in fetal HSC migration and maintenance. Our work paves the way for research into the mechanisms underlying HSC colonization in human fetal BM and provides invaluable resources for future studies on HSC development.

Leptin receptor antagonist attenuates experimental autoimmune thyroiditis in mice by regulating Treg/Th17 cell differentiation.

Leptin has been found to be involved in the development and progression of many autoimmune diseases. As an organ-specific autoimmune disease, the pathogenesis of Hashimoto's thyroiditis has not been fully elucidated. It has been reported that serum leptin level is increased in Hashimoto's thyroiditis, but other studies have not shown any difference. We replicated a mouse model of experimental autoimmune thyroiditis (EAT) with a high-iodine diet and found that injection of the leptin receptor antagonist Allo-aca reduced thyroid follicle destruction and inflammatory cell infiltration in EAT mice, and thyroxine and thyroid autoimmune antibody levels. Further investigation revealed that Allo-aca promotes the differentiation of Treg cells and inhibits the differentiation of Th17 cells. We believe that Allo-aca can alter the differentiation of Treg/Th17 cells by inhibiting the leptin signaling pathway, thereby alleviating thyroid injury in EAT mice. Interfering with the leptin signaling pathway may be a novel new approach to treat treating and ameliorating Hashimoto's thyroiditis.

Whole-exome sequencing expands the roles of novel mutations of organic anion transporting polypeptide, ATP-binding cassette transporter, and receptor genes in intrahepatic cholestasis of pregnancy.

Background: Intrahepatic cholestasis of pregnancy (ICP) is associated with a high incidence of fetal morbidity and mortality. Therefore, revealing the mechanisms involved in ICP and its association with fetal complications is very important. Methods: Here, we used a whole-exome sequencing (WES) approach to detect novel mutations of organic anion transporting polypeptide (OTAP) genes, ATP-binding cassette transporter (ABC) genes, and receptor genes associated with ICP in 249 individuals and 1,029 local control individuals. Two available tools, SIFT and PolyPhen-2, were used to predict protein damage. Protein structuremodeling and comparison between the reference and modified protein structures were conducted by SWISS-MODEL and Chimera 1.14rc software, respectively. Results: A total of 5,583 mutations were identified in 82 genes related to bile acid transporters and receptors, of which 62 were novel mutations. These novel mutations were absent in the 1,029 control individuals and three databases, including the 1,000 Genome Project (1000G_ALL), Exome Aggregation Consortium (ExAC), and Single-Nucleotide Polymorphism Database (dbSNP). We classified the 62 novel loci into two groups (damaging and probably damaging) according to the results of SIFT and PolyPhen-2. Out of the 62 novel mutations, 24 were detected in the damaging group. Of these, five novel possibly pathogenic variants were identified that were located in known functional genes, including ABCB4 (Ile377Asn), ABCB11 (Ala588Pro), ABCC2 (Ile681Lys and Met688Thr), and NR1H4 (Tyr149Ter). Moreover, compared to the wild-type protein structure, ABCC2 Ile681Lys and Met688Thr protein structures showed a slight change in the chemical bond lengths of ATP-ligand binding amino acid side chains. The combined 32 clinical data points indicate that the mutation group had a significantly (p = 0.04) lower level of Cl ions than the wild-type group. Particularly, patients with the 24 novel mutations had higher average values of alanine transaminase (ALT), aspartate transaminase (AST), alkaline phosphatase (ALP), total bile acids (TBA), high-density lipoprotein (HDL), and low-density lipoprotein (LDL) than patients with the 38 novel mutations in the probably damaging group and the local control individuals. Conclusion: The present study provides new insights into the genetic architecture of ICP involving these novel mutations.

Genetic tagging of the adenosine A2A receptor reveals its heterogeneous expression in brain regions.

The adenosine A2A receptor (A2AR), a G protein-coupled receptor, is involved in numerous and varied physiological and pathological processes, including inflammation, immune responses, blood flow, and neurotransmission. Accordingly, it has become an important drug target for the treatment of neuropsychiatric disorders. However, the exact brain distribution of A2AR in regions outside the striatum that display relatively low levels of endogenous A2AR expression has hampered the exploration of A2AR functions under both physiological and pathological conditions. To further study the detailed distribution of the A2AR in low-expression regions, we have generated A2AR knock-in mice in which the 3xHA-2xMyc epitope tag sequence was fused to the C-terminus of A2AR (A2AR-tag mice) via CRISPR/Cas9 technology. Here, using CRISPR/Cas9 technology, we have generated A2AR knock-in mice in which the 3xHA-2xMyc epitope tag sequence was fused to the C-terminus of A2AR (A2AR-tag mice). The A2AR-tag mice exhibited normal locomotor activity and emotional state. Consistent with previous studies, A2AR fluorescence was widely detected in the striatum, nucleus accumbens, and olfactory tubercles, with numerous labeled cells being evident in these regions in the A2AR-tag mouse. Importantly, we also identified the presence of a few but clearly labeled cells in heterogeneous brain regions where A2AR expression has not previously been unambiguously detected, including the lateral septum, hippocampus, amygdala, cerebral cortex, and gigantocellular reticular nucleus. The A2AR-tag mouse represents a novel useful genetic tool for monitoring the expression of A2AR and dissecting its functions in brain regions other than the striatum.