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Interleukin 17D (IL-17D), a pro-inflammatory cytokine, is a signature cytokine of T helper 17 (Th17) cells. However, studies characterizing the functions of IL-17D in teleost are scarce. Therefore, we aimed to characterize the properties of IL-17D in Amphiprion clarkii. We performed spatial and temporal expression, AcIL-17D-mediated antibacterial and inflammatory gene expression, NFκB pathway-related gene expression analyses, and bacterial colony counting and cell protection assays. We found that AcIL-17D contains a 630 bp coding sequence and encodes 210 amino acids. The spatial expression analysis of AcIL-17D in 12 tissues showed ubiquitous expression, with the highest expression in the brain, followed by blood and skin. Temporal expression analysis of AcIL-17D in blood showed upregulated expression at 6 and 24 h (polyinosinic: polycytidylic acid and lipopolysaccharide), 12 h (all stimulants), and 48 h (polyinosinic: polycytidylic acid and Vibrio harveyi). AcIL-17D expression in the blood gradually decreased at later hours in response to all the stimulants. After treatment of fathead minnow (FHM) cells with different recombinant AcIL-17D concentrations, the downstream gene expression analysis showed increased expression of antimicrobial genes in the FHM cells, namely [NK-Lysin (NKL), Hepcidin antimicrobial peptide-1 (HAMP-1), Defensin-β (DEFB1)] and some inflammatory genes such as IL-1β, TNF-α, IL-11, and STAT3. Further nuclear factor κB (NFκB) subunits (NFκB1, NFκB2, RelA, and Rel-B) showed upregulated gene expression at 12 and 24 h. The bacterial colony counting assay using FHM cells showed lower bacterial colony counts in rAcIL-17D-treated cells than in control. Furthermore, the Water-Soluble Tetrazolium Salt (WST -1) assay confirmed the ability of rAcIL-17D in the protection of FHM cells from bacterial infection and conducted the Hoechst 33342 staining upon treatment with rAcIL-17D and rMBP. Therefore, our findings provide important insights into the activation of IL-17D pathway genes in FHM cells, the protective role of AcIL-17D against bacterial infection, and host defense mechanisms in teleost. Show less

Cardiac dysfunction resulting from sepsis causes high morbidity and mortality. Silibinin (SIL) is a secondary metabolite isolated from the seed extract of the milk thistle plant with various properties, including anti-inflammatory, anti-fibrotic, and anti-oxidative activities. This study, for the first time, examined the effects and mechanisms of SIL pretreatment, posttreatment and in combination with classical antibiotics in septic myocardial injury. The survival rate, sepsis score, anal temperature, routine blood parameters, blood biochemical parameters, cardiac function indicators, pathological indicators of myocardial injury, NR1H3 signaling pathway, and several sepsis-related signaling pathways were detected 8 h following cecal ligation and puncture (CLP). Our results showed that SIL pretreatment showed a significant protective effect on sepsis and septic myocardial injury, which was explained by the attenuation of inflammation, inhibition of oxidative stress, improvement of mitochondrial function, regulation of endoplasmic reticulum stress (ERS), and activation of the NR1H3 pathway. SIL posttreatment and the combination of SIL and azithromycin (AZI) showed a certain therapeutic effect. RNA-seq detection further clarified the myocardial protective mechanisms of SIL. Taken together, this study provides a theoretical basis for the application strategy and combination of SIL in septic myocardial injury. Show less

mTOR pathway is known to promote cancer malignancy and influence cancer immunity but is unknown for its role in immune checkpoint inhibitors (ICI) therapy. Using Memorial Sloan-Kettering Cancer Center dataset (MSKCC), we extracted mTOR pathway gene mutations for stepwise Cox regression in 1661 cancer patients received ICI. We associated the mutation of the gene signature resulted from the stepwise Cox regression with the 1661 patients' survival. Other 553 ICI-treated patients were collected from 6 cohorts for validation. We also performed this survival association in patients without ICI treatment from MSKCC as discovery (n = 2244) and The Cancer Genome Atlas (TCGA) as validation (n = 763). Pathway enrichment analysis were performed using transcriptome profiles from TCGA and IMvigor210 trial to investigate the potential mechanism. We identified 8 genes involved in mTOR pathway, including FGFR2, PIK3C3, FGFR4, FGFR1, FGF3, AKT1, mTOR, and RPTOR, resulted from stepwise Cox regression in discovery (n = 1661). In both discovery (n = 1661) and validation (n = 553), the mutation of the 8-gene signature was associated with better survival of the patients treated with ICI, which was independent of tumor mutation burden (TMB) and mainly attributed to the missense mutations. This survival association was not observed in patients without ICI therapy. Intriguingly, the mutation of the 8-gene signature was associated with increased TMB and PD1/PD-L1 expression. Immunologically, pathways involved in anti-tumor immune response were enriched in presence of this mutational signature in mTOR pathway, leading to increased infiltration of immune effector cells (e.g., CD8 + T cells, NK cells, and M1 macrophages), but decreased infiltration of immune inhibitory M2 macrophages. These results suggested that mTOR pathway gene mutations were predictive of better survival upon ICI treatment in multiple cancers, likely by its association with enhanced anti-tumor immunity. Larger studies are warranted to validate our findings. Show less

Heterozygous variants in the leptin-melanocortin pathway are associated with obesity. However, their effect on the long-term outcomes after Roux-en-Y gastric bypass (RYGB) is still unknown. In this matched case-control study, 701 participants from the Mayo Clinic Biobank with a history of RYGB were genotyped. Sixty-three patients had a heterozygous variant in the leptin-melanocortin pathway. After excluding patients with potential confounders, carriers were randomly matched (on sex, age, body mass index [BMI], and years since surgery) with two non-carrier controls. The electronic medical record of carriers and matched non-carriers was reviewed for up to 15 years after RYGB. A total of 50 carriers and 100 matched non-carriers with a history of RYGB were included in the study. Seven different genes (LEPR, PCSK1, POMC, SH2B1, SRC1, MC4R, and SIM1) in the leptin-melanocortin pathway were identified. At the time of surgery, the mean age was 50.8 ± 10.6 years, BMI 45.6 ± 7.3 kg/m Carriers of a heterozygous variant in the leptin-melanocortin pathway have a progressive and significant weight regain in the mid- and long-term after RYGB. Genotyping patients experiencing significant weight regain after RYGB could help implement multidisciplinary and individualized weight loss interventions to improve weight maintenance after surgery. Show less

As one of the common birth defects worldwide, nonsyndromic microtia is a complex disease that results from interactions between environmental and genetic factors. However, the underlying causes of nonsyndromic microtia are currently not well understood. The present study determined transcriptomic and proteomic profiles of auricular cartilage tissues in 10 patients with third-degree nonsyndromic microtia and five control subjects by RNA microarray and tandem mass tag-based quantitative proteomics technology. Relative mRNA and protein abundances were compared and evaluated for their function and putative involvement in nonsyndromic microtia. A total of 3971 differentially expressed genes and 256 differentially expressed proteins were identified. Bioinformatics analysis demonstrated that some of these genes and proteins showed potential associations with nonsyndromic microtia. Thirteen proteins with the same trend at the mRNA level obtained by the integrated analysis were validated by parallel reaction monitoring analysis. Several key genes, namely, Show less

Blood-brain barrier (BBB) injury is involved in the pathogenesis of sepsis-associated encephalopathy. In this study, we used dihydroartemisinin (DHA), a derivative of artemisinin, to treat a cecal ligation and puncture (CLP)-induced mouse sepsis model and a tumour necrosis factor α (TNF-α)-stimulated human cerebral microvessel endothelial cells (hCMEC)/D3 cell line. We found that DHA decreased BBB permeability and increased the expression of the tight junction protein occludin (OCLN) in the CLP model. In hCMEC/D3 cells, DHA decreased TNF-α-induced hyperpermeability and increased the expression of OCLN. DHA also repressed SNAI1 expression in the CLP mouse model and in TNF-α-stimulated hCMEC/D3 cells. These data suggest that DHA protects BBB permeability during sepsis by stimulating the expression of OCLN, by downregulating the expression of the SNAI1 transcription factor. Show less

To investigate the causal role of lipid or apolipoprotein traits in intracerebral hemorrhage (ICH) and determine the effect of lipid-lowering interventions on the disease. Two-sample Mendelian randomization (MR) analyses were conducted to evaluate the associations of high-density lipoprotein cholesterol, low-density lipoprotein cholesterol (LDL-C), triglycerides (TG), apolipoprotein (Apo)B and ApoA1 levels with risks for ICH, and those of LDL-C- (HMGCR, PCSK9, and NPC1L1) and TG-lowering targets (LPL and APOC3) with ICH. Increased levels of ApoB was associated with a decreased risk of overall ICH (OR 0.623, 95% CI 0.413-0.940; p = 0.024) and lobar ICH (OR 0.579, 95% CI 0.342-0.979; p = 0.042). The inverse relationship remained stable in multivariable MR. In addition, elevated TGs showed a causal effect on lobar ICH in multivariable MR (OR 1.600, 95% CI 1.009-2.537; p = 0.046). The LDL-C-reducing genetic variation alleles at or near the HMGCR gene (mimicking the effect of statins) were predicted to increase the overall and deep ICH risk. Additionally, genetic variation at or near the APOC3 gene suggested that genetically reducing the activity of APOC3 (mimicking antisense anti-apoC3 agents) was predicted to decrease lobar ICH. Genetically predicted elevated ApoB may have a protective effect on overall ICH and lobar ICH, whereas elevated TG was associated with a higher risk of lobar ICH conditional on LDL-C and ApoB. MR analysis supports the conclusion that statins may increase the risk of overall and deep ICH independent of their lipid-lowering effect. More specific lipid-lowering targets may end up being the future. ANN NEUROL 2022;92:390-399. Show less

Mutations in VPS13C cause early-onset, autosomal recessive Parkinson's disease (PD). We have established that VPS13C encodes a lipid transfer protein localized to contact sites between the ER and late endosomes/lysosomes. In the current study, we demonstrate that depleting VPS13C in HeLa cells causes an accumulation of lysosomes with an altered lipid profile, including an accumulation of di-22:6-BMP, a biomarker of the PD-associated leucine-rich repeat kinase 2 (LRRK2) G2019S mutation. In addition, the DNA-sensing cGAS-STING pathway, which was recently implicated in PD pathogenesis, is activated in these cells. This activation results from a combination of elevated mitochondrial DNA in the cytosol and a defect in the degradation of activated STING, a lysosome-dependent process. These results suggest a link between ER-lysosome lipid transfer and innate immune activation in a model human cell line and place VPS13C in pathways relevant to PD pathogenesis. Show less

Ameloblastoma is the most common odontogenic tumour and its histomorphological distinction into growth patterns and variants, does not accurately convey information about its biologic aggressiveness. Expression of epithelial mesenchymal transition (EMT) markers, which have been implicated in its etiogenesis, might assist in delineating aggressiveness across variants. This may help in formulating appropriate treatment modalities for its management AIMS: To determine expression of SNAIL/SLUG and ECAD/NCAD in tumour cells in clinical and histological subtypes of ameloblastoma and to establish any association between the immunostaining profile and the biologic behaviour of histologic types of ameloblastoma. This is a retrospective study conducted to evaluate the immunoprofile of selected clinical subtypes and histological variants to EMT factors via immunostaining to SLUG and ECAD/NCAD antibodies. Mean aggregate scores for each antibody per variant was analyzed using ANOVA or Kruskal-Wallis test when appropriate. Agreement between AR and SR regions was correlated using Spearman's correlation co-efficient. A higher staining SLUG intensity in the stellate reticulum (SR) like areas relative to the ameloblast like areas (AR) was observed, without concomitant E-cadherin repression or elimination. However, a direct relationship between SLUG and N-cadherin was observed. Expression of SLUG in the SR like areas can be utilized to predict the biologic behavior of specific clinico-histological variants, however its mechanism via alterations in cadherin switching is equivocal. Show less

Alzheimer's disease (AD) is a neurodegenerative disorder mainly affecting old population. In this study, two Tau overexpressing cell lines (SH-SY5Y/Tau and HEK293/Tau), N2a/SweAPP cell line, and 3× Transgene (APPswe/PS1M146V/TauP301L) mouse primary nerve cell lines were used as AD models to study the activity and molecular mechanism of macelignan, a natural compound extracted from Show less

To explore the expression relationship and significance of long chain non-coding RNA nuclear-enriched abundant transcript 1 (LncRNA NEAT1) and miR-27a-3p in serum and cerebrospinal fluid of patients with Alzheimer's disease (AD). Sixty-six AD patients received by the Department of Neurology of our hospital from October 2019 to September 2021 were gathered, according to the Clinical Dementia Rating Scale (CDR) score, they were grouped into mild group (≤1 point, n = 41) and moderate-to-severe group (> 1 point, n = 25). Another 32 cases of serum and cerebrospinal fluid samples from outpatient physical examination personnel were regarded as the control group. The general materials on all subjects was recorded and cognition was assessed;real-time quantitative PCR was performed to measure the expression levels of miR-27a-3p and NEAT1 in serum and cerebrospinal fluid;enzyme-linked immunosorbent assay was performed to measure the protein levels of β-amyloid precursor protein cleaving enzyme 1 (BACE1), β-amyloid (Aβ) 40 and Aβ42 in cerebrospinal fluid;Spearman's method was performed to analyze the correlation of serum miR-27a-3p and NEAT1 levels with MMSE and MoCA scores;Pearson method was performed to analyze the correlation between serum miR-27a-3p and NEAT1 levels and Aβ deposition standard uptake value ratio (SUVR) and cerebrospinal fluid miR-27a-3p, NEAT1, BACE1, Aβ42 and Aβ40 levels. The MMSE score, MoCA score, serum miR-27a-3p level, cerebrospinal fluid miR-27a-3p, Aβ42 levels and Aβ42/Aβ40 ratio of AD patients in mild group and moderate-to-severe group were all lower than those in the control group, and the moderate-to-severe group were lower than the mild group (all P < 0.05);the serum NEAT1 level, SUVR, and cerebrospinal fluid NEAT1 and BACE1 levels were higher than those in the control group, and the moderate-to-severe group were higher than the mild group (all P < 0.05). Serum NEAT1 level in AD patients was positively correlated with SUVR, cerebrospinal fluid NEAT1 and BACE1 (r = 0.350, 0.606, 0.341, all P < 0.05);serum miR-27a-3p level was positively correlated with cerebrospinal fluid miR-27a-3p level (r = 0.695, P < 0.05), and negatively correlated with SUVR and cerebrospinal fluid BACE1 level (r = - 0.521, - 0.447, both P < 0.05). The expression trends of NEAT1 and miR-27a-3p in the serum and cerebrospinal fluid of AD patients are consistent, the level of NEAT1 is increased, and the level of miR-27a-3p is decreased. The levels of the two are negatively correlated, which is related to the degree of Aβ deposition in the brain of AD patients and is involved in the progression of AD. Show less

Korean pine nut oil (PNO) has been reported to suppress appetite by increasing satiety hormone release. However, previous studies have rendered inconsistent results and there is lack of information on whether dietary Korean PNO affects the expression of satiety hormone receptors and hypothalamic neuropeptides. Therefore, our study sought to evaluate the chronic effects of Korean PNO on the long-term regulation of energy balance. Five-week-old male C57BL/6 mice were fed with control diets containing 10% kcal fat from Korean PNO or soybean oil (SBO) (PC or SC) or high-fat diets (HFDs) containing 35% kcal fat from lard and 10% kcal fat from Korean PNO or SBO (PHFD or SHFD) for 12 weeks. The expression of gastrointestinal satiety hormone receptors, hypothalamic neuropeptides, and genes related to intestinal lipid absorption and adipose lipid metabolism was then measured. There was no difference in the daily food intake between PNO- and SBO-fed mice; however, the PC and PHFD groups accumulated 30% and 18% less fat compared to SC and SHFD, respectively. Korean PNO-fed mice exhibited higher messenger RNA (mRNA) expression of In summary, Korean PNO attenuated body fat accumulation, and appeared to prevent HFD-induced dysregulation of the hypothalamic appetite-suppressing pathway. Show less

We aimed to determine the similarities and differences in the roles of genic and regulatory copy number variations (CNVs) in bipolar disorder (BD), schizophrenia (SCZ), and autism spectrum disorder (ASD). Based on high-resolution CNV data from 8708 Japanese samples, we performed to our knowledge the largest cross-disorder analysis of genic and regulatory CNVs in BD, SCZ, and ASD. In genic CNVs, we found an increased burden of smaller (<100 kb) exonic deletions in BD, which contrasted with the highest burden of larger (>500 kb) exonic CNVs in SCZ/ASD. Pathogenic CNVs linked to neurodevelopmental disorders were significantly associated with the risk for each disorder, but BD and SCZ/ASD differed in terms of the effect size (smaller in BD) and subtype distribution of CNVs linked to neurodevelopmental disorders. We identified 3 synaptic genes (DLG2, PCDH15, and ASTN2) as risk factors for BD. Whereas gene set analysis showed that BD-associated pathways were restricted to chromatin biology, SCZ and ASD involved more extensive and similar pathways. Nevertheless, a correlation analysis of gene set results indicated weak but significant pathway similarities between BD and SCZ or ASD (r = 0.25-0.31). In SCZ and ASD, but not BD, CNVs were significantly enriched in enhancers and promoters in brain tissue. BD and SCZ/ASD differ in terms of CNV burden, characteristics of CNVs linked to neurodevelopmental disorders, and regulatory CNVs. On the other hand, they have shared molecular mechanisms, including chromatin biology. The BD risk genes identified here could provide insight into the pathogenesis of BD. Show less

Echinacoside (ECH), a phenylethanoid glycoside, has protective activity in neurodegenerative disease, including anti-inflammation and antioxidation. However, the effects of ECH in Alzheimer's disease (AD) are not very clear. This present study investigates the role and mechanism of ECH in the pathological process of AD. APP/PS1 mice treated with ECH in 50 mg/kg/day for 3 months. Morris water maze, nesting test, and immunofluorescence staining used to observe whether ECH could improve AD pathology. Western blot used to study the mechanism of ECH improving AD pathology. The results showed that ECH alleviated the memory impairment of APP/PS1 mice by reducing the time of escape latency as well as increasing the times of crossing the platform and rescued the impaired ability to construct nests. In addition, ECH significantly reduced the deposition of senile plaques in the brain and decreased the expression of BACE1 in APP/PS1 mice through activating PI3K/AKT/Nrf2/PPARγ pathway. Furthermore, ECH decreased ROS formation, GP91 and 8-OHdG expression, upregulated the expression of SOD1 and SOD2 as well as activating the PI3K/AKT/Nrf2 signaling pathway. Moreover, ECH inhibited glia cells activation, pro-inflammatory cytokine IL-1β and TNF-α release, NLRP3 inflammasome formation through TXNIP/Trx-1 signaling pathway. In conclusion, this paper reported that ECH improved cognitive function, inhibited oxidative stress, and inflammatory response in AD. Therefore, we suggest that ECH may considered as a potential drug for AD treatment. Show less

Empagliflozin (EMPA) is the first sodium-glucose co-transporter 2 inhibitor to significantly reduce cardiovascular and kidney complications in type 2 diabetes mellitus. Given this, we speculate that EMPA may have the potential to intervene in diabetic retinopathy (DR), which is another diabetes-specific microvascular complication. Db/db mice were treated with EMPA for different periods to observe the retinas and related mechanisms. EMPA effectively balanced body weight and blood glucose levels, mitigated ocular edema and microaneurysm in db/db mice. EMPA significantly inhibited oxidative stress, apoptosis and recovered tight junction in diabetic retinas. MS/MS analyses showed that EMPA suppressed aberrant branched-chain amino acid (BCAAs) accumulation in db/db retinas, which led to the inhibition of the mammalian target of rapamycin activation, downregulation of inflammation, and angiogenic factors, including TNF-ɑ, IL-6, VCAM-1, and VEGF induced by diabetes. Furthermore, branched-chain α-keto acids (BCKAs), which are catabolites of BCAAs, were increased in diabetic retinas and decreased with EMPA application. Moreover, branched-chain ketoacid dehydrogenase kinase (BCKDK) was enhanced, BCKDHA and BCKDHB were decreased in diabetic retinas. This could be reversed by EMPA treatment, thus promoting BCAAs catabolism to decrease BCAAs and BCKAs accumulation in diabetic retinas. The high levels of BCAAs in the plasma and enhanced L-type amino acid transporter 1 (LAT1) were responsible for the high levels of BCAAs in diabetic retinas, which could be inhibited by EMPA. Overall, EMPA could ameliorate DR manifestations. The normalization of BCAAs catabolism and intake may play a role in this process. This study supports EMPA as a protective drug against DR. Show less

Cerebrovascular effects of β-site amyloid precursor protein-cleaving enzyme 1 (BACE1) inactivation have not been systematically studied. In the present study we employed cultured human brain microvascular endothelial cells (BMECs), BACE1-knockout (BACE1 Show less

Histological interpretation of the rare pleomorphic xanthoastrocytoma (PXA) has been the holy grail for treatment options. However, no stand-alone clinical interventions have been developed owing to the lack of gene expression profiling data in PXA/APXA patients. We first time report the comprehensive analyses of the coding as well as long non-coding RNA (lncRNA) signatures of PXA/APXA patients. Several genes such as IGFBP2, NF1, FOS, ERBB2, and lncRNAs such as NEAT1, HOTAIRM1, and GAS5 known to play crucial roles in glioma patients were also deregulated in PXA patients suggesting the commonality in the molecular signatures. PPI network, co-expression, and lncRNA-mRNA interaction studies unraveled hub genes (such as ERBB2, FOS, RPA1) and networks that may play a critical role in PXA biology. The most enriched pathways based on gene profiles were related to TLR, chemokine, MAPK, Rb, and PI3K-Akt signaling pathways. The lncRNA targets were enriched in glucuronidation, adipogenesis, TGF-beta signaling, EGF/EGFR signaling, and cell cycle pathways. Interestingly, several mRNAs like PARVG, and ABI2 were found to be targeted by multiple lncRNAs suggesting a tight control of their levels. Some of the most prominent lncRNA-mRNA pairs were LOC728730: MRPL9, XLOC_l2₀₁₁₉₈₇: ASIC2, lnc-C1QTNF5-1: RNF26. Notably, several lncRNAs such as lnc-CETP-1, lnc-XRCC3-1, lnc-RPL31-1, lnc-USP13-1, and MAPKAPK5-AS1, and genes such as RPA1, NTRK3, and CNRP1 showed strong correlation to the progression-free survival of PXA patients suggesting their potential as novel biomarkers. Overall, the findings of this study may facilitate the development of a new realm of RNA biology in PXA that may have clinical significance in the future. Show less

This study aims to elucidate the role of METTL3 in aggravating the progression of NPC through m Differential expressions of METTL3 in 48 paired NPC tissues and paracancerous tissues were determined by quantitative real-time polymerase chain reaction (qRT-PCR). Its level in NPC patients with different clinical stages and metastatic states was examined. Prognostic potential of METTL3 in NPC patients was assessed by Kaplan-Meier method. After knockdown of METTL3, expression changes of Snail and EMT-related genes, as well as invasive and migratory abilities in SUNE-1 cells were detected. The interaction between Snail with METTL3 and IGF2BP2 was verified by RIP (RNA-Binding Protein Immunoprecipitation) assay. At last, the roles of METTL3/Snail regulatory loop in influencing EMT and metastasis of NPC were clarified. METTL3 was upregulated in NPC tissues than that of paracancerous ones. NPC patients with advanced stage or lymphatic metastasis expressed higher level of METTL3. Kaplan-Meier curves revealed that NPC patients expressing high level of METTL3 suffered worse prognosis. Knockdown of METTL3 downregulated protein levels of Snail and N-cadherin, while E-cadherin was upregulated in SUNE-1 cells. Meanwhile, knockdown of METTL3 inhibited invasive and migratory abilities in NPC cells. RIP assay confirmed the interaction between Snail and METTL3. Besides, knockdown of METTL3 decreased the enrichment abundance of Snail in anti-IGF2BP2. Overexpression of Snail partially reversed the regulatory effects of METTL3 on EMT-related gene expressions and metastatic abilities in NPC. METTL3 is upregulated in NPC, which regulates EMT and metastasis in NPC cells through m Show less

Congenital myasthenic syndrome (CMS) is a clinically and genetically heterogeneous neuromuscular disorder characterized by muscle weakness and caused by mutations in more than 35 different genes. This condition should not be overlooked as a subset of patients with CMS are treatable. However, the diagnosis of CMS is often difficult due to the broad variability in disease severity and course. A five-year-old boy without remarkable family history was born with marked general muscle hypotonia and weakness, respiratory insufficiency, anomalies, and multiple joint contractures. Congenital myopathy was suspected based upon type 1 fiber predominance on muscle biopsy. However, he was diagnosed with CMS at age 4 years when his ptosis and ophthalmoplegia were found to be improved by edrophonium chloride and repetitive nerve stimulation showed attenuation of compound muscle action potentials. An exome sequencing identified a compound heterozygous missense variant of c.737C > T (p.A246V) and a novel intronic insertion c.1166 + 4₁₁₆₆ + 5insAAGCCCACCAC in RAPSN. RT-PCR analysis which showed the skipping of exon 7 in a skeletal muscle sample confirmed that the intronic insertion was pathogenic. His myasthenic symptoms were remarkably improved by pyridostigmine. The patient's diagnosis of CMS was confirmed by exome sequencing, and RT-PCR revealed that the skipping of exon 7 in RAPSN was caused by a novel intronic insertion. The genetic information uncovered in this case should therefore be added to the collection of tools for diagnosing and treating CMS. Show less

Epithelial-mesenchymal transition (EMT) is involved in the pathophysiology of lung cancer (LC) and COPD, and the latter is an important risk factor for LC. We hypothesised that the EMT gene expression profile and signalling cascade may differ in LC patients with COPD from those with no respiratory diseases. In lung tumour specimens obtained through video-assisted thoracoscopic surgery from LC (n=20, control group) and LC-COPD patients (n=30), gene expression (quantitative real-time PCR amplification) of EMT markers Show less

Glucagon-like peptide-1 receptor (GLP-1R) agonists reduce the rates of major cardiovascular events, including myocardial infarction in people with type 2 diabetes, and decrease infarct size while preserving ventricular function in preclinical studies. Nevertheless, the precise cellular sites of GLP-1R expression that mediate the cardioprotective actions of GLP-1 in the setting of ischemic cardiac injury are uncertain. Publicly available single cell RNA sequencing (scRNA-seq) datasets on mouse and human heart cells were analyzed for Glp1r/GLP1R expression. Fluorescent activated cell sorting was used to localize Glp1r expression in cell populations from the mouse heart. The importance of endothelial and hematopoietic cells for the cardioprotective response to liraglutide in the setting of acute myocardial infarction (MI) was determined by inactivating the Glp1r in Tie2+ cell populations. Cardiac gene expression profiles regulated by liraglutide were examined using RNA-seq to interrogate mouse atria and both infarcted and non-infarcted ventricular tissue after acute coronary artery ligation. In mice, cardiac Glp1r mRNA transcripts were exclusively detected in endocardial cells by scRNA-seq. In contrast, analysis of human heart by scRNA-seq localized GLP1R mRNA transcripts to populations of atrial and ventricular cardiomyocytes. Moreover, very low levels of GIPR, GCGR and GLP2R mRNA transcripts were detected in the human heart. Cell sorting and RNA analyses detected cardiac Glp1r expression in endothelial cells (ECs) within the atria and ventricle in the ischemic and non-ischemic mouse heart. Transcriptional responses to liraglutide administration were not evident in wild type mouse ventricles following acute MI, however liraglutide differentially regulated genes important for inflammation, cardiac repair, cell proliferation, and angiogenesis in the left atrium, while reducing circulating levels of IL-6 and KC/GRO within hours of acute MI. Inactivation of the Glp1r within the Tie2+ cell expression domain encompassing ECs revealed normal cardiac structure and function, glucose homeostasis and body weight in Glp1r These findings identify the importance of the murine Tie2+ endothelial cell GLP-1R as a target for the cardioprotective actions of GLP-1R agonists and support the importance of the atrial and ventricular endocardial GLP-1R as key sites of GLP-1 action in the ischemic mouse heart. Hitherto unexplored species-specific differences in cardiac GLP-1R expression challenge the exclusive use of mouse models for understanding the mechanisms of GLP-1 action in the normal and ischemic human heart. Show less

We demonstrated previously that there is a correlation between glucagon-like peptide-1 (GLP-1) single-nucleotide polymorphism (SNP) and bone mineral density in postmenopausal women. Both GLP-1 and glucose-dependent insulinotropic peptide are incretins. The glucose-dependent insulinotropic peptide receptor (GIPR) SNP rs10423928 has been extensively studied. However, it is not clear whether GIPR gene mutations affect bone metabolism. The aim of this study was to investigate the association between rs10423928 and bone mineral density in postmenopausal women in Shanghai. rs10423928 was detected in 884 postmenopausal women in Shanghai, and the correlation between the GIPR SNP and bone mineral density was assessed. The dominant T/T genotype of rs10423928 was found to be related to the bone mineral density of the femoral neck (P = 0.035). Overall, our findings indicate that the dominant T/T genotype of rs10423928 in postmenopausal women is significantly associated with a higher bone mineral density and that the T/T genotype exerts a bone-protective effect. Show less

Obesity is a chronic, multifactorial, relapsing disease. Despite multicomponent lifestyle interventions, including pharmacotherapy, maintaining bodyweight loss is challenging for many people. The pathophysiology of obesity is complex, and currently approved pharmacotherapies only target a few of the many pathways involved; thus, single-targeting agents have limited efficacy. Proglucagon-derived peptides, glucagon, and the incretin hormones glucagon-like peptide-1 (GLP-1) and glucose-dependent insulinotropic polypeptide (GIP), represent attractive targets for managing obesity and metabolic disorders because they may have direct roles in multiple mechanisms including satiety, energy homeostasis, and lipolytic activity. Unimolecular dual and triple agonists targeting glucagon and incretin hormone receptors have been shown to promote bodyweight loss, lower glucose levels, and reduce food intake in animal models of obesity. Multiple dual receptor agonists are in clinical development for the treatment of obesity, including GLP-1/GIP and GLP-1/glucagon receptor agonists. The extent to which glucagon contributes to treatment effects remains to be understood, but it may promote bodyweight loss by reducing food intake, while concomitant GLP-1 receptor agonism ensures normal glucose control. Further research is required to fully understand the molecular mechanisms of action and metabolic effects of both dual and triple receptor agonists. Show less

Cistanche deserticola Ma (cistanche) is a traditional herb with a wide range of therapeutic properties. However, no evidence of cistanche's effect on adipogenesis has been found. The effect of cistanche that promotes the adipogenesis of 3T3-L1 preadipocytes was proved by using MTT spectrophotometry, Nile Red staining, Oil Red O staining and transcriptome sequencing technology. The mRNA level of key transcription factors for adipogenesis such as PPAR, AP2 and LPL were examined by RT-PCR. The results showed that the intracellular lipid content in cistanche treated cells were notably increased when compared with the non-treated cells. Between the differentiation and cistanche treated groups, the expression of adipogenesis related genes such as grow hormone releasing hormone (Ghrp), BCL2/adenovirus E1B interacting protein 3 (Bnip3) and Gastric inhibitory polypeptide receptor (Gipr) were significantly increased. Our findings also verified that cistanche promoted adipogenesis, which was accompanied by up-regulated level of Bnip3 and PPAR. This study could uncover new signaling pathways involved in adipogenesis regulation. Show less

The incretin hormone glucagon-like peptide-1 (GLP-1) has received enormous attention during the past three decades as a therapeutic target for the treatment of obesity and type 2 diabetes. Continuous improvement of the pharmacokinetic profile of GLP-1R agonists, starting from native hormone with a half-life of ~2-3 min to the development of twice daily, daily and even once-weekly drugs highlight the pharmaceutical evolution of GLP-1-based medicines. In contrast to GLP-1, the incretin hormone glucose-dependent insulinotropic polypeptide (GIP) received little attention as a pharmacological target, because of conflicting observations that argue activation or inhibition of the GIP receptor (GIPR) provides beneficial effects on systemic metabolism. Interest in GIPR agonism for the treatment of obesity and diabetes was recently propelled by the clinical success of unimolecular dual-agonists targeting the receptors for GIP and GLP-1, with reported significantly improved body weight and glucose control in patients with obesity and type II diabetes. Here we review the biology and pharmacology of GLP-1 and GIP and discuss recent advances in incretin-based pharmacotherapies. Show less

Gestational diabetes mellitus (GDM) entails a complex underlying pathogenesis, with a specific genetic background and the effect of environmental factors. This study examines the link between a set of single nucleotide polymorphisms (SNPs) associated with diabetes and the development of GDM in pregnant women with different ethnicities, and evaluates its potential modulation with a clinical intervention based on a Mediterranean diet. 2418 women from our hospital-based cohort of pregnant women screened for GDM from January 2015 to November 2017 (the San Carlos Cohort, randomized controlled trial for the prevention of GDM ISRCTN84389045 and real-world study ISRCTN13389832) were assessed for evaluation. Diagnosis of GDM was made according to the International Association of Diabetes and Pregnancy Study Groups (IADPSG) criteria. Genotyping was performed by IPLEX MassARRAY PCR using the Agena platform (Agena Bioscience, SanDiego, CA). 110 SNPs were selected for analysis based on selected literature references. Statistical analyses regarding patients' characteristics were performed in SPSS (Chicago, IL, USA) version 24.0. Genetic association tests were performed using PLINK v.1.9 and 2.0 software. Bioinformatics analysis, with mapping of SNPs was performed using STRING, version 11.5. Quality controls retrieved a total 98 SNPs and 1573 samples, 272 (17.3%) with GDM and 1301 (82.7%) without GDM. 1104 (70.2%) were Caucasian (CAU) and 469 (29.8%) Hispanic (HIS). 415 (26.4%) were from the control group (CG), 418 (26.6%) from the nutritional intervention group (IG) and 740 (47.0%) from the real-world group (RW). 40 SNPs (40.8%) presented some kind of significant association with GDM in at least one of the genetic tests considered. The nutritional intervention presented a significant association with GDM, regardless of the variant considered. In CAU, variants rs4402960, rs7651090, IGF2BP2; rs1387153, rs10830963, MTNR1B; rs17676067, GLP2R; rs1371614, DPYSL5; rs5215, KCNJ1; and rs2293941, PDX1 were significantly associated with an increased risk of GDM, whilst rs780094, GCKR; rs7607980, COBLL1; rs3746750, SLC17A9; rs6048205, FOXA2; rs7041847, rs7034200, rs10814916, GLIS3; rs3783347, WARS; and rs1805087, MTR, were significantly associated with a decreased risk of GDM, In HIS, variants significantly associated with increased risk of GDM were rs9368222, CDKAL1; rs2302593, GIPR; rs10885122, ADRA2A; rs1387153, MTNR1B; rs737288, BACE2; rs1371614, DPYSL5; and rs2293941, PDX1, whilst rs340874, PROX1; rs2943634, IRS1; rs7041847, GLIS3; rs780094, GCKR; rs563694, G6PC2; and rs11605924, CRY2 were significantly associated with decreased risk for GDM. We identify a core set of SNPs in their association with diabetes and GDM in a large cohort of patients from two main ethnicities from a single center. Identification of these genetic variants, even in the setting of a nutritional intervention, deems useful to design preventive and therapeutic strategies. Show less