An interesting and so far unexplained feature of chronic pain in autoimmune disease is the frequent disconnect between pain and inflammation. This is illustrated well in rheumatoid arthritis (RA) where pain in joints (arthralgia) may precede joint inflammation and persist even after successful anti-inflammatory treatment. In the present study, we have addressed the possibility that autoantibodies against citrullinated proteins (ACPA), present in RA, may be directly responsible for the induction of pain, independent of inflammation.

As one of the most essential genome guardians, p53 and its mutants have been suggested associated with many types of cancers. Many p53 mutants function induce unique phenotypes, including carcinogenesis, metastasis, and drug resistance. The p53(R249S) mutation is the most prevalent and specific mutation associated with liver cancer development. Here, we demonstrate the generation of a heterozygous p53(R249S) mutation in the H9 human embryonic stem cell line using TALEN-mediated genome editing. The generated cell line maintains a normal karyotype, a pluripotent state and the in vivo capacity to develop a teratoma containing all three germ layer tissues.

Objectives. Fatty liver disease (FLD) is a major cause of morbidity and mortality worldwide. Dietary cholesterol and alcohol consumption are important risk factors for the progression of FLD, but whether and how alcohol induces more severe FLD with cholesterol ingestion remain unclear. Herein, we mainly used the Lieber-DeCarli diet to establish the FLD mouse model to investigate the synergistic effects of alcohol and cholesterol metabolism on liver damage. The indices of aspartate transaminase (AST), alanine transaminase (ALT), low-density lipoprotein cholesterol (LDL-c), and total cholesterol (TC) levels, inflammation foci, and pathogenesis by hematoxylin and eosin (H&E) and Oil Red O staining revealed that alcohol induces more severe liver damage by influencing cholesterol metabolism, which might be primarily related to the influence of cholesterol absorption, synthesis, and excretion on the liver or small intestine. Moreover, inhibition of absorption of intestinal cholesterol, but not of fat, sucrose, and alcohol, absorption into the body's metabolism by Ezetimibe, significantly improved FLD in rats fed with the high fat-cholesterol-sucrose and alcohol diet. These results showed that alcohol plays an important role in cholesterol metabolism in FLD.

Endothelial cell apoptosis plays an important role in the pathophysiological mechanism of vascular complications in type 2 diabetes mellitus (T2DM). Argirein, a new synthetic compound was demonstrated to inactivate NADPH oxidase to alleviate cardiac dysfunction in T2DM. Here, we investigated whether argirein medication attenuated the vascular dysfunction in T2DM by inhibiting endothelial cell apoptosis which was associated with NADPH oxidase. The rat aortic endothelial cells (RAECs) were incubated with glucose (30 mM) for 48 hour in vitro. It was shown that high glucose significantly increased the protein expression of BAX (Bcl-2 Associated X protein) and Caspase-3 and decreased Bcl2 (B-Cell Leukemia/Lymphoma 2) protein level in RAECs, which was normalized by argirein medication. The annexin V-FITC bound cell percentage and DNA fragments in agarose electrophoresis were markedly suppressed by argirein to confirm the anti-apoptotic property of argirein in RAECs. Furthermore, we found that argirein blocked the endothelin (ET)-1/Nox4 signal-dependent superoxide (O2-.) generation, which regulated endothelial cell apoptosis in RAECs. In vivo, argirein intervention relieved the vasodilatory response to acetylcholine and restored the expressions of Nox4 and BAX in the aorta endothelium of high-fat diet (HFD)-fed rats following streptozocin (STZ) injection. For the first time, we demonstrated that argirein could inhibit vascular endothelial cell apoptosis, which was attributed to blocking ET-1/Nox4 signal-dependent O2- generation in RAECs. This current study revealed the therapeutic effects of argirein to prevent the vascular complication in T2DM through inhibiting endothelial cell apoptosis which was associated with the anti-oxidative property of argirein.

MLL fusion proteins in leukemia induce aberrant transcriptional elongation and associated chromatin perturbations; however, the upstream signaling pathways and activators that recruit or retain MLL oncoproteins at initiated promoters are unknown. Through functional and comparative genomic studies, we identified an essential role for NF-κB signaling in MLL leukemia. Suppression of NF-κB led to robust antileukemia effects that phenocopied loss of functional MLL oncoprotein or associated epigenetic cofactors. The NF-κB subunit RELA occupies promoter regions of crucial MLL target genes and sustains the MLL-dependent leukemia stem cell program. IKK/NF-κB signaling is required for wild-type and fusion MLL protein retention and maintenance of associated histone modifications, providing a molecular rationale for enhanced efficacy in therapeutic targeting of this pathway in MLL leukemias.

Centromeres consist of DNA repeats in many eukaryotes. Non-allelic homologous recombination (HR) between them can result in gross chromosomal rearrangements (GCRs). In fission yeast, Rad51 suppresses isochromosome formation that occurs between inverted repeats in the centromere. However, how the HR enzyme prevents homology-mediated GCRs remains unclear. Here, we provide evidence that Rad51 with the aid of the Swi/Snf-type motor protein Rad54 promotes non-crossover recombination between centromere repeats to prevent isochromosome formation. Mutations in Rad51 and Rad54 epistatically increased the rates of isochromosome formation and chromosome loss. In sharp contrast, these mutations decreased gene conversion between inverted repeats in the centromere. Remarkably, analysis of recombinant DNAs revealed that rad51 and rad54 increase the proportion of crossovers. In the absence of Rad51, deletion of the structure-specific endonuclease Mus81 decreased both crossovers and isochromosomes, while the cdc27/pol32-D1 mutation, which impairs break-induced replication, did not. We propose that Rad51 and Rad54 promote non-crossover recombination between centromere repeats on the same chromatid, thereby suppressing crossover between non-allelic repeats on sister chromatids that leads to chromosomal rearrangements. Furthermore, we found that Rad51 and Rad54 are required for gene silencing in centromeres, suggesting that HR also plays a role in the structure and function of centromeres.

MicroRNAs (miRNAs) are small endogenous non-coding RNAs that regulate post-transcriptional gene expression and are important in many biological processes. Disease-associated miRNAs have been shown to become potential targets for therapeutic intervention. Functions of miRNAs can be inhibited by using antisense oligonucleotides, called anti-miRs, complimentary to the miRNA sequences. Here, we show that systemic delivery of a chemically stabilized anti-miR-122 complexed with interfering nanoparticles (iNOPs) effectively silences the liver-expressed miR-122 in mice. Intravenous administration of 2 mg kg(-1) chemically modified anti-miR-122 complexed with iNOP-7 resulted in 83.2 ± 3.2% specific silencing of miR-122, which was accompanied by regulating gene expression in liver and lowering of plasma cholesterol. The specific silencing of miR-122 was long lasting and did not induce an immune response. Our results demonstrate that iNOPs can successfully deliver anti-miR to specifically target and silence miRNA in clinically acceptable and therapeutically affordable doses.

The generation of reprogrammed induced pluripotent stem cells (iPSCs) from patients with defined genetic disorders holds the promise of increased understanding of the aetiologies of complex diseases and may also facilitate the development of novel therapeutic interventions. We have generated iPSCs from patients with LEOPARD syndrome (an acronym formed from its main features; that is, lentigines, electrocardiographic abnormalities, ocular hypertelorism, pulmonary valve stenosis, abnormal genitalia, retardation of growth and deafness), an autosomal-dominant developmental disorder belonging to a relatively prevalent class of inherited RAS-mitogen-activated protein kinase signalling diseases, which also includes Noonan syndrome, with pleomorphic effects on several tissues and organ systems. The patient-derived cells have a mutation in the PTPN11 gene, which encodes the SHP2 phosphatase. The iPSCs have been extensively characterized and produce multiple differentiated cell lineages. A major disease phenotype in patients with LEOPARD syndrome is hypertrophic cardiomyopathy. We show that in vitro-derived cardiomyocytes from LEOPARD syndrome iPSCs are larger, have a higher degree of sarcomeric organization and preferential localization of NFATC4 in the nucleus when compared with cardiomyocytes derived from human embryonic stem cells or wild-type iPSCs derived from a healthy brother of one of the LEOPARD syndrome patients. These features correlate with a potential hypertrophic state. We also provide molecular insights into signalling pathways that may promote the disease phenotype.

Hypertension is a great global health challenge and it mostly requires drug combination therapy with the various advantages. Metoprolol (MP) and paeoniflorin are both commonly used for the treatment of hypertension. However, whether they exert synergistic effects on antihypertension or not remains unclear, especially on vascular endothelial function.

Post-translational mechanisms regulating cell-matrix adhesion turnover during cell locomotion are not fully elucidated. In this study, we uncovered an essential role of Y118 site-specific tyrosine phosphorylation of paxillin, an adapter protein of focal adhesion complexes, in paxillin recruitment to autophagosomes to trigger turnover of peripheral focal adhesions in human breast cancer cells. We demonstrate that the Rab-7 GTPase is a key upstream regulator of late endosomal sorting of tyrosine118-phosphorylated paxillin, which is subsequently recruited to autophagosomes via the cargo receptor c-Cbl. Essentially, this recruitment involves a direct and selective interaction between Y118-phospho-paxillin, c-Cbl, and LC3 and is independent from c-Cbl E3 ubiquitin ligase activity. Interference with the Rab7-paxillin-autophagy regulatory network using genetic and pharmacological approaches greatly impacted focal adhesion stability, cell locomotion and progression to metastasis using a panel of human breast cancer cells. Together, these results provide novel insights into the requirement of phospho-site specific post-translational mechanism of paxillin for autophagy targeting to regulate cell-matrix adhesion turnover and cell locomotion in breast cancer cells.

The coiled-coil-helix-coiled-coil-helix domain containing 2 (CHCHD2) is overexpressed in several types of cancer. This study aimed to investigate the role of CHCHD2 in hepatocellular carcinoma (HCC). The expression of CHCHD2 in HCC and non-tumorous tissues was detected by immunohistochemistry and Western blot analysis, and the correlation between CHCHD2 expression and clinicopathological features of HCC was analyzed. Furthermore, the proliferation, apoptosis and migration of HepG2 cells with CHCHD2 knockdown were examined. We found that CHCHD2 was upregulated in HCC tissues, and high CHCHD2 expression was associated with poor differentiation, lymph node metastasis, local tissue invasion, high TNM grade of HCC and poor patient survival. Depletion of CHCHD2 led to significantly reduced cell proliferation, increased apoptosis and diminished migratory capacity in HepG2 cells. In addition, HCC tissues had high expression of CD105, a microvessel marker, and HepG2 cells depleted of CHCHD2 had low CD105 expression. In conclusion, CHCHD2 may play an oncogenic role in HCC via promoting tumor cell growth and migration while preventing apoptosis. CHCHD2 is a potential biomarker for poor outcome of HCC patients.

The mechanisms that enable immune evasion at metastatic sites are poorly understood. We show that the Polycomb Repressor Complex 1 (PRC1) drives colonization of the bones and visceral organs in double-negative prostate cancer (DNPC). In vivo genetic screening identifies CCL2 as the top prometastatic gene induced by PRC1. CCL2 governs self-renewal and induces the recruitment of M2-like tumor-associated macrophages and regulatory T cells, thus coordinating metastasis initiation with immune suppression and neoangiogenesis. A catalytic inhibitor of PRC1 cooperates with immune checkpoint therapy to reverse these processes and suppress metastasis in genetically engineered mouse transplantation models of DNPC. These results reveal that PRC1 coordinates stemness with immune evasion and neoangiogenesis and point to the potential clinical utility of targeting PRC1 in DNPC.

Various factors such as ultraviolet rays can cause a continuous threat to our skin, resulting in inflammation or oxidation problems. Ferulic acid (FA), with certain antioxidant and anti-inflammatory properties, is widely used in many cosmetics, even used to treat various diseases in the clinic. In this study, the FA structural skeleton was used to search for FA derivatives. Then, molecular docking, the rule of five, and Veber rules were performed to virtually screen compounds that can bind to proteins with a good drug likeness. DPPH and ABTS were used to evaluate their antioxidant potency and an MTT assay was employed to investigate the toxicities of the compounds, while Griess Reaction System and ELISA were used to judge the concentration variations of NO and different inflammatory factors (TNF-α, IL-1β, and IL-6). Western blotting featured nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) protein expression levels. The trend of the intracellular changes of reactive oxygen species (ROS) was detected by the DCFH-DA method and fluorescence staining. As a result, we found that the ferulic acid derivative S-52372 not only had certain scavenging effects on free radicals in biochemical experiments, but also prevented inflammation and oxidative stress in LPS-stimulated RAW264.7 cells in the cellular environment; intracellular ROS and inflammatory mediators, including iNOS, COX-2, TNF-α, and IL-6, were also suppressed. In a computer prediction, S-52372 owned better water solubility and lower toxicity than FA. This compound deserves further research to find an ideal FA derivative.

Metastasis has emerged to be an important cause for poor prognosis of thyroid carcinoma (TC) and its molecular mechanisms are not fully understood. STRA6 is a multifunctional membrane protein widely expressed in embryonic and adult tissues. The function and mechanism of STRA6 in TC remain elusive.

This study aimed to evaluate the change patterns in corneal intrinsic aberrations and nerve density after cataract surgery in dry eye disease. The preoperative, 1- and 3-month postoperative dry eye-related parameters were obtained by the Oculus keratograph and the ocular surface disease index questionnaire. The corneal intrinsic aberrations were measured using the Pentacam HR system. In vivo confocal microscopy was performed to observe the vortical and peripheral corneal nerves. An artificial intelligence technique run by the deep learning model generated the corneal nerve parameters. Corneal aberrations on the anterior and total corneal surfaces were significantly increased at 1 month compared with the baseline (p < 0.05) but gradually returned to the baseline by 3 months (p > 0.05). However, the change in posterior corneal aberration lasted up to 3 months (p < 0.05). There was a significant decrease in the corneal vortical nerve maximum length and average density after the operation (p < 0.05), and this damage lasted approximately 3 months. The corneal vortical nerve maximum length and average density were negatively correlated with the anterior corneal surface aberrations before and 1 month after the operation (correlation coefficients, CC = −0.26, −0.25, −0.28; all p < 0.05). Corneal vortex provided a unique site to observe long-term corneal nerve injury related to eye dryness. The continuous damage to the corneal vortical nerve may be due to the continuous dry eye state.

Alveolar macrophage (AM) proliferation and self-renewal play an important role in the lung tissue microenvironment. However, the impact of immune cells, especially the neutrophils, on AM homeostasis or function is not well characterized. In this study, we induced in vivo migration of neutrophils into bronchoalveolar lavage (BAL) fluid and lung using CXCL1, and then co-cultured these with AMs in vitro. Neutrophils in the BAL (BAL-neutrophils), rather than neutrophils of bone marrow (BM-neutrophils), were found to inhibit AM proliferation. Analysis of publicly available data showed high heterogeneity of lung neutrophils with distinct molecular signatures of BM- and blood-neutrophils. Unexpectedly, BAL-neutrophils from influenza virus PR8-infected mice (PR8-neutrophils) did not inhibit the proliferation of AMs. Bulk RNA sequencing further revealed that co-culture of AMs with PR8-neutrophils induced IFN-α and -γ responses and inflammatory response, and AMs co-cultured with BAL-neutrophils showed higher expression of metabolism- and ROS-associated genes; in addition, BAL-neutrophils from PR8-infected mice modulated AM polarization and phagocytosis. BAL-neutrophil-mediated suppression of AM proliferation was abrogated by a combination of inhibitors of different neutrophil death pathways. Collectively, our findings suggest that multiple cell death pathways of neutrophils regulate the proliferation of AMs. Targeting neutrophil death may represent a potential therapeutic strategy for improving AM homeostasis during respiratory diseases.

Ras-responsive element-binding protein 1 (Rreb1) is a zinc-finger transcription factor acting downstream of RAS signaling. Rreb1 has been implicated in cancer and Noonan-like RASopathies. However, little is known about its role in mammalian non-disease states. Here, we show that Rreb1 is essential for mouse embryonic development. Loss of Rreb1 led to a reduction in the expression of vasculogenic factors, cardiovascular defects, and embryonic lethality. During gastrulation, the absence of Rreb1 also resulted in the upregulation of cytoskeleton-associated genes, a change in the organization of F-ACTIN and adherens junctions within the pluripotent epiblast, and perturbed epithelial architecture. Moreover, Rreb1 mutant cells ectopically exited the epiblast epithelium through the underlying basement membrane, paralleling cell behaviors observed during metastasis. Thus, disentangling the function of Rreb1 in development should shed light on its role in cancer and other diseases involving loss of epithelial integrity.

In this study, we examined the metabolites from different parts of Acanthopanax senticosus and their role in alleviating damage caused by oxidative stress. We used UHPLC-QTOF-MS to analyze the chemical components in the root, seed, and leaf extracts of A. senticosus. Two multivariate statistical analysis methods-namely, principal component analysis and partial least square discriminant analysis-were used to distinguish the samples obtained from different parts of the plant. Using univariate statistics, 130 different metabolites were screened out. Among these, the relative content of flavonoids and terpenoids was found to be highest in the leaves, the lignin and phenolic acid content was highest in the roots, and the amino acid and phenolic acid levels were highest in seeds. An MTT assay was used to test the anti-H2O2 oxidative damage to PC12 cells in different parts of the sample. Lastly, using Pearson's correlation analysis, various metabolites from different parts of A. senticosus were correlated with their antioxidant effects from the corresponding parts. Fifty-two related different metabolites were found, of which 20 metabolites that were positively correlated to oxidative stress were present at a relatively higher level in the roots, whereas 32 metabolites that were negatively correlated were present at relatively higher levels in the seeds and leaves. The results of this study reveal the distribution characteristics and the antioxidant activity of different metabolites of A. senticosus and provide a reference for the rational development of its medicinal parts.

Cell types have been established during organogenesis based on early mouse embryos. However, our understanding of cell types and molecular mechanisms in the early embryo development of Mongolian sheep has been hampered. This study presents the first comprehensive single-cell transcriptomic characterization at E16 in Ujumqin sheep and Hulunbuir short-tailed sheep. Thirteen major cell types were identified at E16 in Ujumqin sheep, and eight major cell types were identified at E16 in Hulunbuir short-tailed sheep. Function enrichment analysis showed that several pathways were significantly enriched in the TGF-beta signaling pathway, the Hippo signaling pathway, the platelet activation pathway, the riboflavin metabolism pathway, the Wnt signaling pathway, regulation of the actin cytoskeleton, and the insulin signaling pathway in the notochord cluster. Glutathione metabolism, glyoxylate, and dicarboxylate metabolism, the citrate cycle, thyroid hormone synthesis, pyruvate metabolism, cysteine and methionine metabolism, thermogenesis, and the VEGF signaling pathway were significantly enriched in the spinal cord cluster. Steroid biosynthesis, riboflavin metabolism, the cell cycle, the Hippo signaling pathway, the Hedgehog signaling pathway, the FoxO signaling pathway, the JAK-STAT signaling pathway, and the Wnt signaling pathway were significantly enriched in the paraxial mesoderm cluster. The notochord cluster, spinal cord cluster, and paraxial mesoderm cluster were found to be highly associated with tail development. Pseudo-time analysis demonstrated that the mesenchyme can translate to the notochord in Ujumqin sheep. Molecular assays revealed that the Hippo signaling pathway was enriched in Ujumqin sheep. This comprehensive single-cell map revealed previously unrecognized signaling pathways that will further our understanding of the mechanism of short-tailed sheep formation.

Abnormal haemoglobin (Hb) variants result in the most commonly inherited disorders in humans worldwide. In this study, we investigated the molecular epidemiology characteristics of Hb variants, along with associated structural and functional predictions in the Yunnan province population of Southwestern China. A total of 41,933 subjects who sought haemoglobinopathy screening were included. Based on bioinformatics and structural analysis, as well as protein modeling, the pathogenesis and type of Hb genetic mutations were characterized. Among all individuals studied, 328 cases (0.78%) were confirmed as carriers of Hb variants, with 13 cases (0.03%) presenting α-globin variants, 313 (0.75%) β-globin variants, and two δ-globin variants. A total of 19 different mutations were identified, including three novel mutations. In addition, 48 cases of ααCS mutations and 14 cases of Hb H or Hb Bart's were found. The isoelectric point, evolutionary conservation, and genotype-phenotype correlation for these mutations were predicted. Additionally, secondary and tertiary protein structure modeling were performed for three selected mutations. In conclusion, the prevalence of Hb variants in the Yunnan population is much higher than other regions of China. Complete characterization of these Hb variants is essential for generating a rational strategy to control the haemoglobinopathies in this region.