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The present study aims to investigate the alterations of serum proteomic and metabolomic profiles in Chinese patients with severe and active Graves' Orbitopathy (GO). Thirty patients with GO and 30 healthy volunteers were enrolled. The serum concentrations of FT3, FT4, T3, T4, and thyroid-stimulating hormone (TSH) were analyzed, after which TMT labeling-based proteomics and untargeted metabolomics were performed. Metabo- Analyst and Ingenuity Pathway Analysis (IPA) was used for integrated network analysis. A nomogram was established based on the model to explore the disease prediction ability of the identified feature metabolites. One hundred thirteen proteins (19 up-regulated and 94 down-regulated) and 75 metabolites (20 increased and 55 decreased) were significantly altered in GO compared to the control group. By combining the lasso regression, IPA network, and protein-metabolite-disease sub-networks, we extracted feature proteins (CPS1, GP1BA, and COL6A1) and feature metabolites (glycine, glycerol 3-phosphate, and estrone sulfate). The logistic regression analysis revealed that the full model with the prediction factors and three identified feature metabolites had better prediction performance for GO compared to the baseline model. The ROC curve also indicated better prediction performance (AUC = 0.933 vs. 0.789). A new biomarker cluster combined with three blood metabolites with high statistical power can be used to discriminate patients with GO. These findings provide further insights into the pathogenesis, diagnosis, and potential therapeutic targets for this disease. Show less

Nephropathy has become the most common reason for end-stage renal disease worldwide. The progression of end-stage renal disease occurs caused by decreased glomerular filtration rate, damage to capillaries in renal glomeruli or a higher risk of cardiovascular morbidity and mortality in diabetic patients. The involvement of mechanism in the development of nephropathy via generation of AGEs, the elevation of growth factors, altered hemodynamic and metabolic factors, inflammatory mediators, oxidative stress and dyslipidaemia. The prevalence of chronic kidney disease in India will rise from 3.7 million in 1990 to 7.63 million in 2020 becoming the main cause of mortality and morbidity. The pathogenesis of nephropathy mediates by various molecules that cause alterations in the structure and function of the kidney like growth factors, endothelins, transforming growth factor (TGF-β), and Angiotensin-converting enzymes (ACE), fibronectin and proinflammatory cytokines, mast cells and dyslipidemia. Growth factors like VEGF, IGFs, PDGF, EGFR and TGF-β contribute to excessive extracellular matrix accumulation, together with thickening of the glomerular and tubular basement membranes and an increase in the mesangial matrix, leading to glomerulosclerosis and tubulointerstitial fibrosis. Oxidative stress and inflammation factors like TNF-α, IL-1 and IL-6 are hypothesized to play a role in the development of pathological changes in nephropathy like renal hyperfiltration and hypertrophy, thickening of the glomerular basement membrane (GBM), glomerular lesion and tubulointerstitial fibrosis. Dyslipidemia is involved in the progression of nephropathy by impaired action of lipoprotein lipase, lecithincholesterol acyltransferase (LCAT) and cholesteryl ester transferase protein (CETP) resulting in the increased level of LDL-C, Triglyceride level and decrease HDL-C that enhance macrophage infiltration, excessive extracellular matrix production and accelerate inflammation with the development of proteinuria. Interruption in the RAS, oxidative stress and dyslipidemia have yielded much better results in terms of reno-protection and progression of nephropathy. In this review, we would focus on various factors that have been shown to contribute to renal injury in many experimental models of nephropathy. Show less

Endometriosis is an inflammatory disease characterized by the presence of ectopic endometrial tissue, immune cell dysfunction and abnormal cytokine secretion. In addition to immunological factors, genetic variations that influence endometriosis severity and cytokine expression levels play important roles in the pathogenesis of this disease. Interleukin-12 (IL-12), specifically its p40 subunit encoded by IL-12B gene and the interleukin-12 receptor β1 (IL-12Rβ2) chain of its receptor, as well as interleukin-27 (IL-27) are important in the establishment of endometriosis. So, in this study, we measured IL-12 and IL-27 serum levels and investigated the possible links between IL-12B rs3212227, IL-12Rβ2 rs3790565 and IL-27 rs153109 polymorphisms and the risk of developing endometriosis in a group of Iranian women. In this case-control study, 162 endometriosis patients and 151 healthy women were included and tested for the aforementioned polymorphisms using the polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) technique. The enzyme-linked immunosorbent assay (ELISA) method was also used to measure IL-12 and IL-27 serum levels. Although there was no statistically significant association between the genotypes and alleles of the studied polymorphisms and the development of endometriosis in general, the AA genotype of IL-12B rs3212227 showed a significant association with uterine endometriosis when compared to AC+CC genotypes (p = .04, CI = 0.270-0.988, OR = 0.517). Indeed, the AA genotype of the IL-12B rs3212227 single nucleotide polymorphism (SNP) may be linked with a lower risk of developing uterine endometriosis. There was no significant difference in IL-27 levels between the two studied groups (p = .49), and IL-12 levels were undetectable in both groups. In conclusion, the AA genotype of IL-12B rs3212227 might be associated with a decreased risk of uterine involvement in endometriosis patients. Show less

Due to aging of the population, bone frailty is dramatically increasing worldwide. Although some therapeutic options exist, they do not fully protect or prevent against the occurrence of new fractures. All current drugs approved for the treatment of bone fragility target bone mass. However, bone resistance to fracture is not solely due to bone mass but relies also on bone extracellular matrix (ECM) material properties, i.e., the quality of the bone matrix component. Here, we introduce the first-in-class unimolecular dual glucose-dependent insulinotropic polypeptide/glucagon-like peptide-2 (GIP/GLP-2) analogue, GL-0001, that activates simultaneously the glucose-dependent insulinotropic polypeptide receptor (GIPr) and the glucagon-like peptide-2 receptor (GLP-2r). GL-0001 acts synergistically through a cyclic adenosine monophosphate-lysyl oxidase pathway to enhance collagen maturity. Furthermore, bilateral ovariectomy was performed in 32 BALB/c mice at 12 weeks of age prior to random allocation to either saline, dual GIP/GLP-2 analogues (GL-0001 or GL-0007) or zoledronic acid groups (n = 8/group). Treatment with dual GIP/GLP-2 analogues was initiated 4 weeks later for 8 weeks. At the organ level, GL-0001 modified biomechanical parameters by increasing ultimate load, postyield displacement, and energy-to-fracture of cortical bone. GL-0001 also prevented excess trabecular bone degradation at the appendicular skeleton and enhanced bone ECM material properties in cortical bone through a reduction of the mineral-to-matrix ratio and augmentation in enzymatic collagen cross-linking. These results demonstrate that targeting bone ECM material properties is a viable option to enhance bone strength and opens an innovative pathway for the treatment of patients suffering from bone fragility. © 2023 The Authors. Journal of Bone and Mineral Research published by Wiley Periodicals LLC on behalf of American Society for Bone and Mineral Research (ASBMR). Show less

Apolipoprotein A-IV (ApoA-IV) plays a role in satiation and serum lipid transport. In diet-induced obesity (DIO) C57BL/6J mice, ApoA-IV deficiency induced in ApoA-IV-/-knock-out (KO mice) resulted in increased bodyweight, insulin resistance (IR) and plasma free fatty acid (FFA), which was partially reversed by stable ApoA-IV-green fluorescent protein (KO-A4-GFP) transfection in KO mice. DIO KO mice exhibited increased M1 macrophages in epididymal white adipose tissue (eWAT) as well as in the blood. Based on RNA-sequencing analyses, cytokine-cytokine receptor interactions, T cell and B cell receptors, and especially IL-17 and TNF-α, were up-regulated in eWAT of DIO ApoA-IV KO compared with WT mice. Supplemented ApoA-IV suppressed lipopolysaccharide (LPS)-induced IKK and JNK phosphorylation in Raw264.7 macrophage cell culture assays. When the culture medium was supplemented to 3T3-L1 adipocytes they exhibited an increased sensitivity to insulin. ApoA-IV protects against obesity-associated metabolic inflammation mainly through suppression in M1 macrophages of eWAT, IL17-IKK and IL17-JNK activity. Show less

Allergic rhinitis (AR) is a nasal allergic disease mainly mediated by IgE, and the immune response is the pathological basis of AR pathogenesis. Regulatory T cells (Treg) has been confirmed to be involved in the occurrence of AR. IL-27 mediates inflammatory responses and allergic symptoms in AR by promoting Tregs and related factors. Our study aimed to explore the correlation between serum interleukin 27 (IL-27) and Treg associated cytokines in the pathogenesis of AR. Based on the inclusion and exclusion criteria, 69 participants with AR and 50 healthy participants were selected. Their IL-27, IL-10, and TGF-β1 levels were estimated by ELISA. Receiver operating characteristics curve (ROC) was performed to demonstrate the diagnostic efficiency of IL-27 in AR. Pearson correlation analysis was used for correlation analysis. IL-27 in AR patients statistically decreased compared to the control group. Consistently, IL-27 were also negatively correlated with the clinical severity of AR patients. Treg related cytokines including IL-10 and TGF-β1 in AR patients was statistically decreased. In addition, the IL-10 and TGF-β1expressions were positively correlated with IL-27 in AR patients. IL-27 was statistically down-regulated in patients with AR, which is related to insufficient Treg function. Restoring the expression of IL-27 may become a novel approach to treat AR. Show less

CPS1, the rate-limiting enzyme that controls the first reaction of the urea cycle, is responsible for converting toxic ammonia into non-toxic urea in mammals. While disruption of the functions of CPS1 leads to elevated ammonia and nerve damage in the body, mainly manifested as urea cycle disorder. Moreover, accumulating evidence has recently revealed that CPS1 is involved in a variety of human diseases, including CPS1D, cardiovascular disease, cancers, and others. In particular, CPS1 expression varies among cancers, being overexpressed in some cancers and downregulated in others, suggesting that CPS1 may be a promising cancer therapeutic target. In addition, some small-molecule inhibitors of CPS1 have been reported, which have not been confirmed experimentally in malignancies, meaning their future role is far from certain. In this review, we describe the structure and function of CPS1, highlight its important roles in various human diseases, and further discuss the potential diagnostic and therapeutic implications of small molecule compounds targeting CPS1. Show less

Despite many advances in treatment over the past few years, the poor 5-year survival rate and high recurrence rate of gastric cancer (GC) remain unsatisfactory. As the most abundant epigenetic modification in the eukaryotic mRNA, N6-methyladenosine (m Show less

Large-scale genome-wide association studies (GWASs) on bipolar disorder (BD) have implicated the involvement of the fatty acid desaturase (FADS) locus. These enzymes (FADS1 and FADS2) are involved in the metabolism of eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA), which are thought to potentially benefit patients with mood disorders. To model reductions in the activity of FADS1/2 affected by the susceptibility alleles, we generated mutant mice heterozygously lacking both Fads1/2 genes. We measured wheel-running activity over six months and observed bipolar swings in activity, including hyperactivity and hypoactivity. The hyperactivity episodes, in which activity was far above the norm, usually lasted half a day; mice manifested significantly shorter immobility times on the behavioral despair test performed during these episodes. The hypoactivity episodes, which lasted for several weeks, were accompanied by abnormal circadian rhythms and a marked decrease in wheel running, a spontaneous behavior associated with motivation and reward systems. We comprehensively examined lipid composition in the brain and found that levels of certain lipids were significantly altered between wild-type and the heterozygous mutant mice, but no changes were consistent with both sexes and either DHA or EPA was not altered. However, supplementation with DHA or a mixture of DHA and EPA prevented these episodic behavioral changes. Here we propose that heterozygous Fads1/2 knockout mice are a model of BD with robust constitutive, face, and predictive validity, as administration of the mood stabilizer lithium was also effective. This GWAS-based model helps to clarify how lipids and their metabolisms are involved in the pathogenesis and treatment of BD. Show less

To investigate the role of genetic testing in patients with idiopathic atrioventricular conduction disease requiring pacemaker (PM) implantation before the age of 50 years. All consecutive PM implantations in Southern Switzerland between 2010 and 2019 were evaluated. Inclusion criteria were: (i) age at the time of PM implantation: < 50 years; (ii) atrioventricular block (AVB) of unknown aetiology. Study population was investigated by ajmaline challenge and echocardiographic assessment over time. Genetic testing was performed using next-generation sequencing panel, containing 174 genes associated to inherited cardiac diseases, and Sanger sequencing confirmation of suspected variants with clinical implication. Of 2510 patients who underwent PM implantation, 15 (0.6%) were young adults (median age: 44 years, male predominance) presenting with advanced AVB of unknown origin. The average incidence of idiopathic AVB computed over the 2010-2019 time window was 0.7 per 100 000 persons per year (95% CI 0.4-1.2). Most of patients (67%) presented with specific genetic findings (pathogenic variant) or variants of uncertain significance (VUS). A pathogenic variant of PKP2 gene was found in one patient (6.7%) with no overt structural cardiac abnormalities. A VUS of TRPM4, MYBPC3, SCN5A, KCNE1, LMNA, GJA5 genes was found in other nine cases (60%). Of these, three unrelated patients (20%) presented the same heterozygous missense variant c.2531G > A p.(Gly844Asp) in TRPM4 gene. Diagnostic re-assessment over time led to a diagnosis of Brugada syndrome and long-QT syndrome in two patients (13%). No cardiac events occurred during a median follow-up of 72 months. Idiopathic AVB in adults younger than 50 years is a very rare condition with an incidence of 0.7 per 100 000 persons/year. Systematic investigations, including genetic testing and ajmaline challenge, can lead to the achievement of a specific diagnosis in up to 20% of patients. Heterozygous missense variant c.2531G > A p.(Gly844Asp) in TRPM4 gene was found in an additional 20% of unrelated patients, suggesting possible association of the variant with the disease. Show less

Alzheimer's Disease (AD) is a complex and progressive neurodegenerative disease, and the most common cause of dementia usually occurs due to old age. Production and accumulation of amyloid-β peptide (Aβ) represent the major pathological event of the disease. The formation of amyloid- β results due to proteolytic cleavage of amyloid precursor protein (APP) by beta-site amyloid precursor protein cleaving enzyme (BACE1) shown as the amyloid hypothesis, a prevalent theory for AD pathogenesis. Thus, BACE1 represents a novel target to decrease cerebral Aβ concentration and slow down the disease's progression. The structure-based drug design approach led to a wide variety of small molecules with the mechanism of action centered around inhibition of β-secretase protease (BACE1), which are shown to have drug-like properties and reduce brain Aβ levels. Based on transition state isosteres, BACE1 inhibitors can largely be classified as peptidomimetics and non-peptidomimetics. The subclasses of the two categories have been covered with different scaffolds like statin, norstatin, carbinamine, hydroxyethylene, hydroxyethylamine, acyl guanidine, 2- aminopyridine, aminoimidazole, aminohydantoin, aminothiazoline, aminooxazoline, aminoquinoline, piperazine-based. Among these small molecules, those who fulfilled general requirements for a drug aimed at the central nervous system (CNS) and selectivity over other aspartyl proteases reached the final pipeline of clinical trials. Here, in this review, we summarize the journey of BACE1 inhibitors through different practices of drug design development, Structural Activity Relationship (SAR), and other inhibitor candidates that are currently in clinical trials as BACE1 inhibitors. Show less

Beta amyloid cleaving enzyme 1 (BACE1) is largely expressed by neurons and is the sole β-secretase for initiating the production of neuronal β-amyloid peptides (Aβ). To fully understand the physiological functions of neuronal BACE1, we used mouse genetic approach coupled with unbiased single nucleus RNA sequencing (snRNAseq) to investigate how targeted deletion of Bace1 in neurons, driven by Thy-1-Cre recombinase, would affect functions in the nervous system. Our transcriptome results revealed that BACE1 is essential for maturation of neural precursor cells and oligodendrocytes in mice. RNA velocity analysis confirmed deficit in the trajectory of neuroblasts in reaching the immature granule neuron state in young Bace1fl/fl; Thy1-cre mice. Further analysis of differential gene expression indicated changes in genes important for SNARE signaling, tight junction signaling, synaptogenesis and insulin secretion pathways. Morphological studies revealed a hypomyelination in Bace1fl/fl;Thy1-cre sciatic nerves, but no detectable myelination changes in the corpus callosum, despite clear reduction in myelination proteins in the brain. Functional studies showed reduction in long-term potential, defects in synaptogenesis and learning behavioral. Altogether, our results show that neuronal BACE1 is critical for optimal development of central and peripheral nervous system, and inhibition of neuronal BACE1 will result in deficits in synaptic functions and cognitive behaviors. Show less

Thyroid-associated ophthalmopathy (TAO) is a chronic autoimmune disease. The interleukin-12 (IL-12) family includes IL-12, IL-23, IL-27, and IL-35, all of which play important roles in autoimmunity. Thus far, the relationship between IL-12, IL-27, and IL-35 and the TAO has not been evaluated. Seventy-five serum samples from patients with TAO were collected. Serum samples from 90 healthy controls (HC), 55 patients with Graves' disease (GD), 38 patients with uveitis (UV), 17 patients with Sjogren's syndrome (SS), and 65 patients with rheumatoid arthritis (RA) were collected as controls. The associations between IL-27, IL-35, IL-12, and other clinical parameters were analyzed. Elevated serum levels of IL-27/IL-35 and decreased serum IL-12 levels were observed in TAO patients compared to those in HC (p < 0.001). For HC, we observed good diagnostic ability to predict TAO (area under the curve = 0.74, 0.78, and 0.78, for IL-27, IL-35, and IL-12, respectively). For other autoimmune diseases, IL-27, IL-35, and IL-12 had the ability to discriminate between UV, RA, and SS (area under the curve = 0.80, 0.83, and 0.85 for IL-27; 0.52, 0.69, and 0.67 for IL-35). The positive detection rates of IL-12 were significantly lower in the TAO group than in the UV and RA groups (p = 0.002, 0.01). IL-12, IL-27, and IL-35 have the potential as biomarkers for TAO. Show less

Aggregates of β-amyloid peptide are found to occur in brains of AD patients and are formed upon sequential cleavage of the amyloid precursor protein by BACE1 and γ-secretase. Strategies inhibiting either peptide aggregation or the rate limiting enzyme BACE1 have been in demand for its implication in AD therapeutics. The present study is undertaken to mine compounds with dual ability. In this context, some natural compounds that were already predicted as BACE1 inhibitors by our group, were further tested for their activity as aggregation inhibitors. A pharmacophore model built with known antiamyloidogenic compounds was then applied for screening the natural compounds previously predicted as BACE1 inhibitors. Subsequently experimental validation by Thioflavin-T and Aβ-GFP assay filtered four compounds genistein, syringetin, tamarixetin and ZINC53276039. Out of them, ZINC53276039 showed promising antiamyloidogenic activity to act as a potent inhibitor of aggregation. Interestingly, our previous study revealed syringetin and ZINC53276039 to be good BACE1 inhibitors while tamarixetin to be a moderate BACE1 inhibitor. These good to moderate BACE1 inhibitors with moderate to reasonable antiamyloidogenic activity might show potency in reducing the amyloid load of AD brains. Show less

The medial prefrontal cortex (mPFC), one of the most vulnerable brain regions in Alzheimer's disease (AD), plays a critical role in cognition. Leucine-rich repeat and immunoglobulin-like domain-containing nogo receptor-interacting protein-1 (LINGO-1) negatively affects nerve growth in the central nervous system; however, its role in the pathological damage to the mPFC remains to be studied in AD. In this study, an anti-LINGO-1 antibody was administered to 10-month-old APP/PS1 mice, and behavioral tests, stereological methods, immunohistochemistry and immunofluorescence were used to answer this question. Our results revealed that LINGO-1 was highly expressed in the neurons of the mPFC of AD mice, and the anti-LINGO-1 antibody improved prefrontal cortex-related function and reduced the protein level of LINGO-1, atrophy of the volume, Aβ deposition and massive losses of synapses and neurons in the mPFC of AD mice. Antagonizing LINGO-1 could effectively alleviate the pathological damage in the mPFC of AD mice, which might be an important structural basis for improving prefrontal cortex-related function. Abnormal expression of LINGO-1 in the mPFC may be one of the key targets of AD, and the effect initiated by the anti-LINGO-1 antibody may provide an important basis in the search for drugs for the prevention and treatment of AD. Show less

Emerging evidence has shown lncRNAs play important roles in signaling pathways involved in colorectal cancer (CRC) carcinogenesis. However, only a few functional lncRNAs have been extensively researched, especially in CRC-related signaling pathways. Looking for novel candidate regulators of CRC incidence and progression, using available RNA-seq and microarray datasets, LINC00963 was introduced as a bona fide oncogenic-lncRNA. Consistently, RT-qPCR results showed that LINC00963 was up-regulated in CRC tissues. However, our attempt to amplify the full-length lncRNA from cDNA resulted in the discovery of two novel variants (LINC00963-v2 & LINC00963-v3) that surprisingly, were downregulated in CRC tissues, detected by RT-qPCR. Overexpression of LINC00963-v2/-v3 in HCT116 and SW480 cells resulted in downregulation of the major oncogenes and upregulation of the main tumor suppressor genes involved in PI3K and Wnt signaling, verified through RT-qPCR, western blotting, and TOPFlash assays. Mechanistic studies revealed that LINC00963-v2/-v3 exert their effect on PI3K and Wnt signaling through sponging miR-10a-5p, miR-143-3p, miR-217, and miR-512-3p, which in turn these miRNAs are fine-regulators of PTEN, APC1, and Axin1 tumor suppressor genes verified by dual-luciferase assay and RT-qPCR. At cellular levels, LINC00963-v2/-v3 overexpression suppressed cell proliferation, viability, and migration while increasing the apoptosis of CRC cell lines, detected by PI flow cytometry, colony formation, MTT, RT-qPCR, wound-healing, Transwell, AnnexinV-PE/7AAD, caspase3/7 activity assays, and Hoechst/PI-AO/EB staining. Overall, our results indicate that LINC00963-v2 & -v3 are novel tumor suppressor ceRNAs that attenuate the PI3K and Wnt pathways during CRC incidence and these lncRNAs may serve as potential targets for CRC therapy. Show less

Angiopoietin-like protein 4 (ANGPTL4) is produced in chronic or acute inflammation. Although ANGPTL4 increases in the periodontal ligament fibroblasts during hypoxia, the involvement and role of ANGPTL4 in periodontitis have not been elucidated. In this study, we investigated whether ligature-induced experimental periodontitis and/or Porphyromonas gingivalis lipopolysaccharides (Pg-LPS) would upregulate ANGPTL4 expression and whether ANGPTL4 would somehow involve in the expression of matrix metalloproteinases (MMPs) which are key molecules in the process of periodontal tissue destruction. Experimental periodontitis was induced in 6-week-old male Sprague-Dawley rats by placing a nylon suture around the neck of the maxillary second molar. Two weeks after the induction of periodontitis, the periodontal tissue was excised and analyzed by histological/immunohistochemical staining and gene expression analyses. Human gingival fibroblasts (hGFs) were stimulated with Pg-LPS. The gene expression of ANGPTLs and receptors involved in ANGPTL4 recognition were observed. We also confirmed the changes in gene expression of MMPs upon stimulation with human ANGPTL4. Furthermore, we downregulated ANGPTL4 expression by short interfering RNA in hGFs and investigated the effect of Pg-LPS on MMP production. Induction of periodontitis significantly increased the expression of ANGPTL4 in the gingiva. Pg-LPS significantly increased the gene and protein expression of ANGPTL4 in hGFs but not the gene expression of other ANGPTLs or ANGPTL receptors. Recombinant human ANGPTL4 significantly increased MMP13 gene expression in hGFs. We also confirmed that MMP13 expression was increased in the gingiva during experimental periodontitis. Pg-LPS induced MMP13 gene expression in hGFs. These results suggest the pivotal role of ANGPTL4 in periodontitis. Periodontitis increases ANGPTL4 expression in the gingiva, further suggesting that increased ANGPTL4 may be a factor involved in enhancing MMP13 expression. Show less

Large tumor suppressor kinase 1 (LATS1), one of the predominant components of the Hippo pathway, has been characterized as a key player controlling the proliferation and invasion of cancer cells, including gastric cancer (GC) cells. However, the mechanism by which the functional stability of LATS1 is modulated has yet to be elucidated. Online prediction tools, immunohistochemistry and western blotting assays were used to explore the expression of WW domain-containing E3 ubiquitin ligase 2 (WWP2) in GC cells and tissues. Gain- and loss-of-function assays, as well as rescue experiments were performed to determine the role of the WWP2-LATS1 axis in cell proliferation and invasion. Additionally, the mechanisms involving WWP2 and LATS1 were assessed by coimmunoprecipitation (Co-IP), immunofluorescence, cycloheximide and in vivo ubiquitination assays. Our results demonstrate a specific interaction between LATS1 and WWP2. WWP2 was markedly upregulated and correlated with disease progression and a poor prognosis in GC patients. Moreover, ectopic WWP2 expression facilitated the proliferation, migration and invasion of GC cells. Mechanistically, WWP2 interacts with LATS1, resulting in its ubiquitination and subsequent degradation, leading to increased transcriptional activity of YAP1. Importantly, LATS1 depletion abolished the suppressive effects of WWP2 knockdown on GC cells. Furthermore, WWP2 silencing attenuated tumor growth by regulating the Hippo-YAP1 pathway in vivo. Our results define the WWP2-LATS1 axis as a critical regulatory mechanism of the Hippo-YAP1 pathway that promotes GC development and progression. Video Abstract. Show less

Chronic kidney disease (CKD) is challenging to reverse and its treatment options are limited. Yishen-Qingli-Huoxue Formula (YQHF) is an effective treatment Chinese formula for CKD, as verified by clinical randomized controlled trial. However, the correlative YQHF therapeutic mechanisms are still unknown. The current study aimed to investigate the potential anti-renal fibrosis effects of YQHF as well as the underlying mechanism. After affirming the curative effects of YQHF on adenine-induced CKD rats, Masson staining, immunohistochemistry, and ELISA were used to assess the effects of YQHF on renal fibrosis. Subsequently, metabolomics and transcriptomics analyses were conducted to clarify the potential mechanisms. Furthermore, high performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS), molecular docking analysis and in vitro experiments were used to verify final mechanism of anti-fibrosis. Our results demonstrated that YQHF could improve renal morphology, decrease blood urea nitrogen (BUN), serum creatinine (Scr), and increase body weight gain of model rats. Masson staining, immunohistochemistry of collagen I, fibronectin (FN), α-smooth muscle actin (α-SMA), vimentin and E-cadherin showed that YQHF delayed CKD progression by alleviating renal fibrosis, and the expression of fibrotic factors smoc2 and cdh11 were obviously suppressed by YQHF. Metabolomic and transcriptomic measures discovered that indoxyl sulfate might be a crucial factor inducing renal fibrosis, and the antagonistic effect of YQHF on renal fibrosis may be exerted via AhR/snai1 signaling. Subsequently, western blot and immunohistochemical experiments revealed YQHF indeed inhibited AhR/snai1 signaling in adenine-induced renal fibrosis of CKD rat, which confirmed previous results. In addition, molecular docking and in vitro experiments further supported this conclusion, in which astilbin, the main compound identified YQHF, was certified to exert a significant effect on AhR. Our findings showed that YQHF can effectively treat CKD by antagonizing renal fibrosis, the potential mechanisms were relating with the regulation on AhR/snai1 signaling. Show less

Alzheimer's disease (AD) is a neurodegenerative, progressive, and fatal disorder characterized by marked atrophy of the cerebral cortex and loss of basal forebrain cholinergic neurons. The main pathological features of AD are related to neuronal degeneration and include extracellular deposition of amyloid beta plaques (Aβ plaques), intracellular formation of neurofibrillary tangles (NFTs), and neuroinflammation. So far, drugs used to treat AD have symptomatic and palliative pharmacological effects, disappearing with continued use due to neuron degeneration and death. Therefore, there are still problems with an effective drug for treating AD. Few approaches evaluate the action of natural products other than alkaloids on the molecular targets of β-amyloid protein (Aβ protein) and/or tau protein, which are important targets for developing neuroprotective drugs that will effectively contribute to finding a prophylactic drug for AD. This review gathers and categorizes classes of natural products, excluding alkaloids, which in silico analysis (molecular docking) and in vitro and/or in vivo assays can inhibit the BACE1 and GSK-3β enzymes involved in AD. Show less

The large HDL particles generated by administration of cholesteryl ester transfer protein inhibitors (CETPi) remain poorly characterized, despite their potential importance in the routing of cholesterol to the liver for excretion, which is the last step of the reverse cholesterol transport. Thus, the effects of the CETPi dalcetrapib and anacetrapib on HDL particle composition were studied in rabbits and humans. The association of rabbit HDL to the LDL receptor (LDLr) in vitro was also evaluated. New Zealand White rabbits receiving atorvastatin were treated with dalcetrapib or anacetrapib. A subset of patients from the dal-PLAQUE-2 study treated with dalcetrapib or placebo were also studied. In rabbits, dalcetrapib and anacetrapib increased HDL-C by more than 58% (P < 0.01) and in turn raised large apo E-containing HDL by 66% (P < 0.001) and 59% (P < 0.01), respectively. Additionally, HDL from CETPi-treated rabbits competed with human LDL for binding to the LDLr on HepG2 cells more than control HDL (P < 0.01). In humans, dalcetrapib increased concentrations of large HDL particles (+69%, P < 0.001) and apo B-depleted plasma apo E (+24%, P < 0.001), leading to the formation of apo E-containing HDL (+47%, P < 0.001) devoid of apo A-I. Overall, in rabbits and humans, CETPi increased large apo E-containing HDL particle concentration, which can interact with hepatic LDLr. The catabolism of these particles may depend on an adequate level of LDLr to contribute to reverse cholesterol transport. Show less

Fatty acid desaturase (FADS) genetic polymorphisms are strongly correlated with the risk of dyslipidemia and cardiovascular disease. In this study, we examined the impact of FADS1 and FADS2 genetic variants on plasma lipid status, and assessed interactions between FADS genetic polymorphisms and plasma n-3/n-6 fatty acids regarding lipid status within a population of 816 Taiwanese patients with type 2 diabetes. Selected tag single-nucleotide polymorphisms (FADS1 rs174546 [T/C]; FADS2 rs174602 [A/G] and rs2072114 [A/G]) were genotyped (n = 816). The distribution of genotypes were compared with reports publicly available in the Genome Aggregation Database for East Asian populations (https://gnomad.broadinstitute.org). In the subgroup of patients not taking lipid-lowering medications (n = 192), we observed that the G allele of FADS2 rs174602 was statistically significantly correlated with lower low-density lipoprotein cholesterol (LDL-C) concentrations (P = 0.001), whereas the G allele of rs2072114 was marginally associated with LDL-C concentrations (P = 0.091). Using a general linear model adjusted for confounding factors, statistically significant interactions (P = 0.016) between single-nucleotide polymorphisms in rs2072114 and a low alpha-linolenic acid (18:3n-3)/linoleic acid (18:2n-6) ratio; the G allele correlated with lower LDL-C levels among individuals with a low alpha-linolenic acid/linoleic acid ratio. Interaction between rs174602 single-nucleotide polymorphisms and low alpha-linolenic acid/linoleic acid values on LDL-C was only marginally significant (P = 0.063). Our results show the role of n-3/n-6 dietary polyunsaturated fatty acids in modifying the effects of genetic susceptibility on lipoprotein concentrations in patients with type 2 diabetes. Our findings highlight the potential of interventions with dietary polyunsaturated fatty acids regarding developing individualized prevention strategies for type 2 diabetes presenting with co-occurring dyslipidemia and cardiovascular diseases. Show less

Tumor radiation resistance is the main obstacle to effective radiation therapy for patients with hepatocellular carcinoma (HCC). We identified the role of urea cycle key enzyme carbamoyl phosphate synthetase 1 (CPS1) in radioresistance of HCC and explored its mechanism, aiming to provide a novel radiosensitization strategy for the CPS1-deficiency HCC subtype. The expression of CPS1 was measured by western blot and immunohistochemistry. Cell growth assay, EdU assay, cell apoptosis assay, cell cycle assay, clone formation assay, and subcutaneous tumor assay were performed to explore the relationship between CPS1 and radioresistance of HCC cells. Lipid metabonomic analysis was used for investigating the effects of CPS1 on lipid synthesis of HCC cells. RNA sequencing and coimmunoprecipitation assay were carried out to reveal the mechanism of CPS1 participating in the regulation of HCC radiation therapy resistance. Furthermore, 10074-G5, the specific inhibitor of c-Myc, was administered to HCC cells to investigate the role of c-Myc in CPS1-deficiency HCC cells. We found that urea cycle key enzyme CPS1 was frequently lower in human HCC samples and positively associated with the patient's prognosis. Functionally, the present study proved that CPS1 depletion could accelerate the development of HCC and induce radiation resistance of HCC in vitro and in vivo, and deficiency of CPS1 promoted the synthesis of some lipid molecules. Regarding the mechanism, we uncovered that inhibition of CPS1 upregulated CyclinA2 and CyclinD1 by stabilizing oncoprotein c-Myc at the posttranscriptional level and generated radioresistance of HCC cells. Moreover, inactivation of c-Myc using 10074-G5, a specific c-Myc inhibitor, could partially attenuate the proliferation and radioresistance induced by depletion of CPS1. Our results recapitulated that silencing CPS1 could promote HCC progression and radioresistance via c-Myc stability mediated by the ubiquitin-proteasome system, suggesting that targeting c-Myc in CPS1-deficiency HCC subtype may be a valuable radiosensitization strategy in the treatment of HCC. Show less

Platelet concentrate (PC) is an alternative therapy to treat mastitis in dairy cattle and is an alternative treatment for reproduction problems such as endometritis. Unfortunately, double-centrifugation processing methods described are time-consuming, require specialized laboratory equipment, and are usually done in a heterologous way, which risks herd health. To overcome this limitation, we evaluated single-step bovine PC processing methods readily applicable to a farm setting using an autologous conditioned plasma (ACP) production system. We investigated the hematologic findings, cytokines, and growth factors of the obtained PC samples. Autologous conditioned plasma was prepared using whole blood (WB) from 4 cows (group 1) using single-step centrifugation and 16 different processing methods. The 2 protocols that yielded the highest ratio of platelet to white blood cell (WBC) concentration were ACP-1 [720 × g (2,200 rpm), 5 min] and ACP-2 [929 × g (2,500 rpm), 3 min]. They were subsequently reproduced and compared using WB from 8 cows (group 2). Hematologic findings were quantified, IL-1β (cytokine) and growth factors [platelet-derived growth factor (PDGF), transforming growth factor (TGF)-β, bovine fibroblast growth factor (b-FGF)] were measured, and enrichment factors were compared between samples and processing methods. Hematological characteristics and platelet enrichment varied markedly among tested protocols and all were statistically different from WB. Protocol ACP-2 resulted in significantly greater platelet enrichment (mean 169% of WB) than ACP-1 (125% of WB). We found no significant difference between the 2 ACP preparation protocols with regard to leukocyte reduction (7.53-9.75% WBC compared with WB) or growth factor enrichment (124-125% PDGF, 95-100% TGF-β, 102-104% b-FGF, and 56-74% IL-1β compared with WB). In conclusion, both ACP protocols yielded a platelet concentration shown to promote healing for clinical applications in cattle, and the ACP-2 protocol resulted in a greater degree of platelet enrichment. Therefore, this protocol could be used for ACP production for clinical applications in cattle. Show less

Increasing evidence supports the involvement of the peripheral immune system in the pathogenesis of Alzheimer's disease (AD). In the present study, we found that B lymphocytes could mitigate beta-Amyloid (Aβ) pathology and memory impairments in a transgenic AD mouse model. Specifically, in young 5 × FAD mice, we evidenced increased B cells in the frontal cortex and meningeal tissues; depletion of mature B cells aggravated these mice's Aβ load and memory deficits. The increased B cells produced more interleukin-35 (IL-35) in the front cortex. We further found IL-35 neutralization exacerbated Aβ pathology, while injecting IL-35 mitigated Aβ load and cognitive dysfunction in 5 × FAD mice with or without mature B cell deficiency. Mechanistically, IL-35 inhibited neuronal BACE1 transcription through modulating the SOCS1/STAT1 pathway, and reduced Aβ production accordingly. Reanalysis of the single-cell RNA sequencing data from blood samples of AD patients suggested an increased population of IL-35-producing B cells. Together, the present study revealed a novel effect of B lymphocyte-derived IL-35 on inhibiting Aβ production in the frontal cortex, which may serve as a potential target for future AD treatment. Show less

The melanocortin-4 receptor (MC4R) is a centrally expressed, class A GPCR that plays a key role in the regulation of appetite and food intake. Deficiencies in MC4R signaling result in hyperphagia and increased body mass in humans. Antagonism of MC4R signaling has the potential to mitigate decreased appetite and body weight loss in the setting of anorexia or cachexia due to underlying disease. Herein, we report on the identification of a series of orally bioavailable, small-molecule MC4R antagonists using a focused hit identification effort and the optimization of these antagonists to provide clinical candidate Show less

Beta-site secretase (BACE1) catalyzes the cleavage of amyloid precursor protein (APP), which process ultimately lead to plaque deposition in the brain of Alzheimer's disease (AD). Thus, accurate monitor of BACE1 activity is essential to screen inhibitors for AD treatment. This study develops a sensitive electrochemical assay for probing BACE1 activity based on silver nanoparticles (AgNPs) and tyrosine conjugation as tags and a marking method, respectively. An APP segment is firstly immobilized on aminated microplate reactor. Cytosine (C) rich sequence-templated AgNPs/Zr-based metal-organic framework (MOF) composite is modified by phenol groups, and then the prepared tag (ph-AgNPs@MOF) is captured in microplate surface by the conjugation reaction of phenolic groups between tyrosine and tag. After cleavage by BACE1, the solution containing ph-AgNPs@MOF tags is transferred to the screen-printed graphene electrode (SPGE) surface for voltammetric detection of AgNP signal. This sensitive detection for BACE1 provided an excellent linear relationship between 1 to 200 pM with a detection limit of 0.8 pM. Furthermore, this electrochemical assay is successfully applied for screening of BACE1 inhibitors. This strategy is also verified to be used for evaluation of BACE1 in serum samples. Show less

Human epicardial adipose tissue (EAT) plays a crucial role in the development and progression of coronary artery disease, atrial fibrillation, and heart failure. Microscopically, EAT is composed of adipocytes, nerve tissues, inflammatory, stromovascular, and immune cells. Epicardial adipose tissue is a white adipose tissue, albeit it also has brown fat-like or beige fat-like features. No muscle fascia divides EAT and myocardium; this allows a direct interaction and crosstalk between the epicardial fat and the myocardium. Thus, it might be a therapeutic target for pharmaceutical compounds acting on G-protein-coupled receptors, such as those for glucose-dependent insulinotropic polypeptide (GIP), glucagon (GCG), and glucagon-like peptide-1 (GLP-1), whose selective stimulation with innovative drugs has demonstrated beneficial cardiovascular effects. The precise mechanism of these novel drugs and their tissue and cellular target(s) need to be better understood. We evaluate whether human EAT expresses GIP, GCG, and GLP-1 receptors and whether their presence is related to EAT transcriptome. We also investigated protein expression and cell-type localization specifically for GIP receptor (GIPR) and glucagon receptor (GCGR). Epicardial adipose tissue samples were collected from 33 patients affected by cardiovascular diseases undergoing open heart surgery (90.9% males, age 67.2 ± 10.5 years mean ± SD). Microarray and immunohistochemistry analyses were performed. Microarray analysis showed that GIPR and GCGR messenger ribonucleic acids (mRNAs) are expressed in EAT, beyond confirming the previously found GLP-1 [3776 ± 1377 arbitrary unit (A.U.), 17.77 ± 14.91 A.U., and 3.41 ± 2.27 A.U., respectively]. The immunohistochemical analysis consistently indicates that GIPR and GCGR are expressed in EAT, mainly in macrophages, isolated, and in crown-like structures. In contrast, only some mature adipocytes of different sizes showed cytoplasmic immunostaining, similar to endothelial cells and pericytes in the capillaries and pre-capillary vascular structures. Notably, EAT GIPR is statistically associated with the low expression of genes involved in free fatty acid (FFA) oxidation and transport and those promoting FFA biosynthesis and adipogenesis (P < 0.01). Epicardial adipose tissue GCGR, in turn, is related to genes involved in FFA transport, mitochondrial fatty acid oxidation, and white-to-brown adipocyte differentiation, in addition to genes involved in the reduction of fatty acid biosynthesis and adipogenesis (P < 0.01). Having reported the expression of the GLP-1 receptor previously, here, we showed that GIPR and GCGR similarly present at mRNA and protein levels in human EAT, particularly in macrophages and partially adipocytes, suggesting these G-protein-coupled receptors as pharmacological targets on the ongoing innovative drugs, which seem cardiometabolically healthy well beyond their effects on glucose and body weight. Show less