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BACKGROUND: Adolescent idiopathic scoliosis (AIS), the predominant genetic-influenced scoliosis, results in spinal deformities without vertebral malformations. However, the molecular aetiology of AIS remains unclear. METHODS: Using genome/exome sequencing, we studied 368 patients with severe AIS (Cobb angle >40°) and 3794 controls from a Han Chinese cohort. We performed gene-based and pathway-based weighted rare variant association tests to assess the mutational burden of genes and established biological pathways. Differential expression analysis of muscle tissues from 14 patients with AIS and 15 controls was served for validation. RESULTS: SLC16A8, a lactate transporter linked to retinal glucose metabolism, was identified as a novel severe AIS-associated gene (p=3.08E-06, false discovery rate=0.009). Most AIS cases with deleterious SLC16A8 variants demonstrated early onset high myopia preceding scoliosis. Pathway-based burden test also revealed a significant enrichment in multiple carbohydrate metabolism pathways, especially galactose metabolism. Patients with deleterious variants in these genes demonstrated a significantly larger spinal curve. Genes related to catabolic processes and nutrient response showed divergent expression between AIS cases and controls, reinforcing our genomic findings. CONCLUSION: This study uncovers the pivotal role of genetic variants in carbohydrate metabolism in the development of AIS, unveiling new insights into its aetiology and potential treatment.
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Metabolismo de los Hidratos de Carbono , Escoliosis , Humanos , Escoliosis/genética , Escoliosis/patología , Adolescente , Femenino , Masculino , Metabolismo de los Hidratos de Carbono/genética , Predisposición Genética a la Enfermedad , Niño , Secuenciación del Exoma , Transportadores de Ácidos Monocarboxílicos/genética , Estudios de Casos y Controles , Estudios de Asociación Genética , MutaciónRESUMEN
OBJECTIVE: Long QT syndrome type 7 (Andersen-Tawil syndrome, ATS), which is caused by KCNJ2 gene mutation, often leads to ventricular arrhythmia, periodic paralysis and skeletal malformations. The development, differentiation and electrophysiological maturation of cardiomyocytes (CMs) changes promote the pathophysiology of Long QT syndrome type 7(LQT7). We aimed to specifically reproduce the ATS disease phenotype and study the pathogenic mechanism. METHODS AND RESULTS: We established a cardiac cell model derived from human induced pluripotent stem cells (hiPSCs) to the phenotypes and electrophysiological function, and the establishment of a human myocardial cell model that specifically reproduces the symptoms of ATS provides a reliable platform for exploring the mechanism of this disease or potential drugs. The spontaneous pulsation rate of myocardial cells in the mutation group was significantly lower than that in the repair CRISPR group, the action potential duration was prolonged, and the Kir2.1 current of the inward rectifier potassium ion channel was decreased, which is consistent with the clinical symptoms of ATS patients. Only ZNF528, a chromatin-accessible TF related to pathogenicity, was continuously regulated beginning from the cardiac mesodermal precursor cell stage (day 4), and continued to be expressed at low levels, which was identified by WGCNA method and verified with ATAC-seq data in the mutation group. Subsequently, it indicated that seven pathways were downregulated (all p < 0.05) by used single sample Gene Set Enrichment Analysis to evaluate the overall regulation of potassium-related pathways enriched in the transcriptome and proteome of late mature CMs. Among them, the three pathways (GO: 0008076, GO: 1990573 and GO: 0030007) containing the mutated gene KCNJ2 is involved that are related to the whole process by which a potassium ion enters the cell via the inward rectifier potassium channel to exert its effect were inhibited. The other four pathways are related to regulation of the potassium transmembrane pathway and sodium:potassium exchange ATPase (p < 0.05). ZNF528 small interfering (si)-RNA was applied to hiPSC-derived cardiomyocytes for CRISPR group to explore changes in potassium ion currents and growth and development related target protein levels that affect disease phenotype. Three consistently downregulated proteins (KCNJ2, CTTN and ATP1B1) associated with pathogenicity were verificated through correlation and intersection analysis. CONCLUSION: This study uncovers TFs and target proteins related to electrophysiology and developmental pathogenicity in ATS myocardial cells, obtaining novel targets for potential therapeutic candidate development that does not rely on gene editing.
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Síndrome de Andersen , Células Madre Pluripotentes Inducidas , Humanos , Síndrome de Andersen/diagnóstico , Síndrome de Andersen/genética , Cromatina/metabolismo , Transcriptoma , Mutación/genética , Miocitos Cardíacos/metabolismo , Potasio/metabolismoRESUMEN
BACKGROUND: The indications and outcome of surgery for Acute type A aortic dissection (ATAAD) in elderly patients are still debated, especially when they were above 80 years old. Case presentation: This report describes the case of an octogenarian patient with ATAAD who underwent total arch replacement (TAR) combined with stented elephant trunk (SET) implantation. CONCLUSION: Emergent surgery should be performed on the ATAAD octogenarians without serious preoperative complications. Acceptable outcomes could be received by total arch replacement combined with SET implantation.
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Aneurisma de la Aorta Torácica , Disección Aórtica , Implantación de Prótesis Vascular , Anciano , Anciano de 80 o más Años , Disección Aórtica/diagnóstico , Disección Aórtica/cirugía , Aneurisma de la Aorta Torácica/diagnóstico , Aneurisma de la Aorta Torácica/cirugía , Humanos , Octogenarios , Resultado del TratamientoRESUMEN
Thoracic aortic aneurysm and dissection (TAAD) is a life-threatening disease and currently there is no pharmacological therapy. Sympathetic nerve overactivity plays an important role in the development of TAAD. Sympathetic innervation is mainly controlled by nerve growth factor (NGF, a key neural chemoattractant) and semaphoring 3A (Sema3A, a key neural chemorepellent), while the roles of these two factors in aortic sympathetic innervation and especially TAAD are unknown. We hypothesized that genetically manipulating the NGF/Sema3A ratio by the Ngf -driven Sema3a expression approach may reduce aortic sympathetic nerve innervation and mitigate TAAD progression. A mouse strain of Ngf gene-driven Sema3a expression (namely NgfSema3a/Sema3a mouse) was established by inserting the 2A-Sema3A expression frame to the Ngf terminating codon using CRISPR/Cas9 technology. TAAD was induced by ß-aminopropionitrile monofumarate (BAPN) both in NgfSema3a/Sema3a mice and wild type (WT) littermates. Contrary to our expectation, the BAPN-induced TAAD was severer in NgfSema3a/Sema3a mice than in wild-type (WT) mice. In addition, NgfSema3a/Sema3a mice showed higher aortic sympathetic innervation, inflammation and extracellular matrix degradation than the WT mice after BAPN treatment. The aortic vascular smooth muscle cells isolated from NgfSema3a/Sema3a mice and pretreated with BAPN in vivo for two weeks showed stronger capabilities of proliferation and migration than that from the WT mice. We conclude that the strategy of Ngf -driven Sema3a expression cannot suppress but worsens the BAPN-induced TAAD. By investigating the aortic phenotype of NgfSema3a/Sema3a mouse strain, we unexpectedly find a path to exacerbate BAPN-induced TAAD which might be useful in future TAAD studies.
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Aneurisma de la Aorta Torácica , Disección Aórtica , Azidas , Desoxiglucosa , Animales , Ratones , Aminopropionitrilo/efectos adversos , Aneurisma de la Aorta Torácica/genética , Aneurisma de la Aorta Torácica/inducido químicamente , Aneurisma de la Aorta Torácica/metabolismo , Desoxiglucosa/análogos & derivados , Modelos Animales de Enfermedad , Factor de Crecimiento Nervioso/genética , Factor de Crecimiento Nervioso/efectos adversos , Semaforina-3A/genéticaRESUMEN
Congenital vertebral malformation, affecting 0.13-0.50 per 1000 live births, has an immense locus heterogeneity and complex genetic architecture. In this study, we analyze exome/genome sequencing data from 873 probands with congenital vertebral malformation and 3794 control individuals. Clinical interpretation identifies Mendelian etiologies in 12.0% of the probands and reveals a muscle-related disease mechanism. Gene-based burden test of ultra-rare variants identifies risk genes with large effect sizes (ITPR2, TBX6, TPO, H6PD, and SEC24B). To further investigate the biological relevance of the genetic association signals, we perform single-nucleus RNAseq on human embryonic spines. The burden test signals are enriched in the notochord at early developmental stages and myoblast/myocytes at late stages, highlighting their critical roles in the developing spine. Our work provides insights into the developmental biology of the human spine and the pathogenesis of spine malformation.
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Anomalías Musculoesqueléticas , Columna Vertebral , Humanos , Columna Vertebral/anomalías , Anomalías Musculoesqueléticas/genética , Alelos , Exoma , Proteínas de Dominio T Box/genéticaRESUMEN
PURPOSE: Hepatocellular carcinoma (HCC) is one of the most common cancers and a leading cause of death worldwide. Accurate prognosis prediction tools are urgently needed. While the use of circulating tumor cells (CTCs) as prognostic prediction tool has a clear potential. METHODS: We established a comprehensive, negative enrichment-based strategy for CTCs analysis in patients with HCC, involving identification of epithelial CTCs (E-CTCs) and mesenchymal CTCs (M-CTCs) through specific biomarker. This strategy was performed in 127 HCC cases, 21 nonmalignant liver disease (NMLD) patients and 42 health control to analyze the relevance between CTCs and tumor recurrence. RESULTS: The total CTC number and M-CTC percent were positively correlated with tumor malignancy and high recurrence risk. Individually, preoperative total CTC number and M-CTC percent could robustly distinguish relapse cases from those with no relapse, with sensitivity of 80.95% and 90.48%, specificity of 74.12% and 84.71%, respectively. Levels of preoperative total CTC number and M-CTC percent can both be regarded as independent risk factors for HCC with early recurrence (P = 0.0053, P < 0.0001), and are both significantly correlated with worse recurrence-free survival (RFS) (log rank P < 0.0001; HR 7.78, 95% CI = 3.59-16.87; log rank P < 0.0001; HR 24.4, 95% CI = 8.67-68.77). The levels of total CTC number and M-CTC number had higher effectiveness than alpha fetal protein (AFP) in HCC longitudinal supervision (77.78% vs 88.89% vs 22.22%). CONCLUSION: Preoperative and postoperative CTCs with higher effectiveness than AFP in prognosis prediction and recurrence supervision, indicating that CTCs could work as the biomarker for HCC clinical management.
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Carcinoma Hepatocelular , Neoplasias Hepáticas , Células Neoplásicas Circulantes , Humanos , Carcinoma Hepatocelular/patología , Células Neoplásicas Circulantes/patología , Neoplasias Hepáticas/patología , alfa-Fetoproteínas , Recurrencia Local de Neoplasia , Pronóstico , Biomarcadores , Biomarcadores de Tumor/metabolismoRESUMEN
OBJECTIVE: This study evaluated the early and late outcomes of non-total aortic arch replacement for acute Stanford A aortic dissection. METHODS: 131 cases of acute Stanford Type A aortic dissection with no rupture admitted to our hospital from January 2016 to December 2019 were selected for non-total aortic arch replacement. According to different surgical methods, 51 patients with tear-oriented ascending/hemiarch replacement were included in Group A, and 80 patients who underwent total arch replacement surgery were enrolled in Group B. The perioperative indicators, 30-day mortality rate, and the incidence of postoperative complications were compared between the two groups, and the survival rate of patients were compared by follow-up after discharge. RESULTS: The cardiopulmonary bypass time, cardiac perfusion time, invasive ventilation and ICU hospitalization in Group A were critically shorter than those in Group B (P<0.05). The incidence of transient cerebral dysfunction in Group A was substantially lower than that in Group B (P<0.05). The difference of comparison in perioperative mortality, incidence of permanent neurological dysfunction, and incidence of acute kidney and liver damage between the two groups was statistically insignificant (P>0.05). In addition, the two groups had statistically insignificant difference in survival during postoperative follow-up (P>0.05). CONCLUSION: For acute Stanford type A aortic dissection without rupture in aortic arch, the non-total aortic arch replacement has simple surgical method with high perioperative safety and long-term efficacy that similar to total arch replacement.
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Nutritional microenvironment determines the specification of progenitor cells, and lipid availability was found to modulate osteogenesis in skeletal progenitors. Here, we investigated the implications of lipid scarcity in the osteogenic differentiation of bone marrow mesenchymal stromal cells (BMSCs) and the role of low-density lipoprotein receptor-related protein 5 (LRP5), a co-receptor transducing canonical Wnt/beta-catenin signals, in BMSC lipid uptake during osteogenesis. The osteogenic differentiation of murine BMSCs was suppressed by lipid scarcity and partially rescued by additional fatty acid treatment with oleate. The enhancement of osteogenesis by oleate was found to be dosage-dependent, along with the enhanced activation of beta-catenin and Wnt target genes. Conditional knockout (CKO) of Lrp5 gene in murine mesenchymal lineage using Lrp5 fl/fl ;Prrx1-cre mice led to decreased bone quality and altered fat distribution in vivo. After Lrp5 ablation using adenoviral Cre-recombinase, the accumulation of lipid droplets in BMSC cytoplasm was significantly reduced, and the osteogenesis of BMSCs was suppressed. Moreover, the impaired osteogenesis due to either lipid scarcity or Lrp5 ablation could be rescued by recombinant Wnt3a protein, indicating that the osteogenesis induced by Wnt/beta-catenin signaling was independent of LRP5-mediated lipid uptake. In conclusion, lipid scarcity suppresses BMSC osteogenic differentiation. LRP5 plays a role in the uptake of lipids in BMSCs and therefore mediates osteogenic specification.
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Genetic perturbations in nicotinamide adenine dinucleotide de novo (NAD) synthesis pathway predispose individuals to congenital birth defects. The NADSYN1 encodes the final enzyme in the de novo NAD synthesis pathway and, therefore, plays an important role in NAD metabolism and organ embryogenesis. Biallelic mutations in the NADSYN1 gene have been reported to be causative of congenital organ defects known as VCRL syndrome (Vertebral-Cardiac-Renal-Limb syndrome). Here, we analyzed the genetic variants in NADSYN1 in an exome-sequenced cohort consisting of patients with congenital vertebral malformations (CVMs). A total number of eight variants in NADSYN1, including two truncating variants and six missense variants, were identified in nine unrelated patients. All enrolled patients presented multiple organ defects, with the involvement of either the heart, kidney, limbs, or liver, as well as intraspinal deformities. An in vitro assay using COS-7 cells demonstrated either significantly reduced protein levels or disrupted enzymatic activity of the identified variants. Our findings demonstrated that functional variants in NADSYN1 were involved in the complex genetic etiology of CVMs and provided further evidence for the causative NADSYN1 variants in congenital NAD Deficiency Disorder.
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Ligasas de Carbono-Nitrógeno con Glutamina como Donante de Amida-N/genética , Enfermedades de la Columna Vertebral/congénito , Enfermedades de la Columna Vertebral/genética , Columna Vertebral/anomalías , Secuencia de Aminoácidos , Animales , Células COS , Ligasas de Carbono-Nitrógeno con Glutamina como Donante de Amida-N/química , Chlorocebus aethiops , Estudios de Cohortes , Humanos , Mutación , Alineación de Secuencia , Secuenciación del ExomaRESUMEN
Obscurin participates in the development of striated muscles and maintenance of the functional sarcoplasmic reticulum. However, the role of obscurin in arrhythmogenic right ventricular cardiomyopathy (ARVC) is not well understood. We aimed to study the novel obscurin mutations in the pathogenesis of ARVC and the underlying mechanisms. Methods: We generated induced pluripotent stem cells (iPSC) through retroviral reprogramming of peripheral blood mononuclear cells isolated from a 46-year-old female diagnosed with ARVC, carrying a mutation in OBSCN. The cells differentiated into functional iPSC-based cardiomyocytes (iPSC-CMs), whose phenotype was determined by transmission electron microscopy, electrophysiological description, immunofluorescence staining, and Oil Red O staining. Molecular characterization was performed by bioinformatic analyses, and identification by quantitative real-time polymerase chain reaction (qRT-PCR) and Western blotting. Results: ARVC-iPSC-CMs mutation in OBSCN showed significant accumulation of lipids, increased pleomorphism, irregular Z-bands, and increased L type calcium currents. Functional enrichment analysis identified pathways involved in focal adhesion and structure formation; the adipocytokines and PPAR signaling pathways were also activated in the ARVC group. Moreover, our results from ultra-high-resolution microscopy, qRT-PCR and Western blotting confirmed that the mutant OBSCN protein and its anchor protein, Ank1.5, showed structural disorder and decreased expression, but there was increased expression of junctional protein N-Cadherin. Further analysis revealed the gene expression of other desmosomal proteins in ARVC-iPSC-CMs was also decreased but some adipogenesis pathway-related proteins (PPARγ, C/EBPα, and FABP4) were increased. Conclusion: A novel frameshift mutation in OBSCN caused phenotypic alteration accompanied by disrupted localization and decreased expression of its anchoring protein Ank1.5. Furthermore, there was an accumulation of lipids with an increase in fatty fibrosis area and myocardial structural disorder, possibly leading to dysrhythmia in calcium channel-related myocardial contraction. These observations suggested the possibility of attenuating ARVC progression by therapeutic modulation of OBSCN expression.
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Displasia Ventricular Derecha Arritmogénica/genética , Calcio/metabolismo , Miocardio/patología , Miocitos Cardíacos/patología , Proteínas Serina-Treonina Quinasas/genética , Factores de Intercambio de Guanina Nucleótido Rho/genética , Animales , Displasia Ventricular Derecha Arritmogénica/patología , Células Cultivadas , Análisis Mutacional de ADN , Femenino , Fibroblastos , Fibrosis , Mutación del Sistema de Lectura , Humanos , Células Madre Pluripotentes Inducidas , Cariotipificación , Masculino , Ratones , Persona de Mediana Edad , Miocardio/citología , Técnicas de Placa-Clamp , Cultivo Primario de Células , Proteínas Serina-Treonina Quinasas/metabolismo , Factores de Intercambio de Guanina Nucleótido Rho/metabolismo , Secuenciación del ExomaRESUMEN
Chemical immunosuppressants have been widely used for the treatment of systemic lupus erythematosus (SLE). However, these small chemical drugs suffer from poor solubility, short circulating half-life and adverse side effects. One of the most effective strategies to extend the circulating time is loading drugs into nanocarriers to form nanomedicines, which is of particular interest for the treatment of cancer and viral diseases but has seldom been applied to autoimmune disorders. Herein, we successfully developed an easy but general drug delivery platform based on the new biocompatible polyhydroxyalkanoate (PHA) terpolymer poly(3-hydroxybutyrate-co-3-hydroxyvalerate-co-3-hydroxyhexanoate) (PHBVHHx). In this proof of concept study, we loaded the PHBVHHx nanocarrier with the immunosuppressant azathioprine (AZA) for SLE therapy for the first time. The AZA-PHA nanoparticles possessed â¼30% cytotoxicity and slow clearance from the kidneys. In a murine SLE model, AZA-PHA nanoparticles exhibited superior therapeutic efficacy to AZA and AZA-polylactic acid (PLA) nanoparticles without appreciable toxicity. This delivery system may provide a new and general platform for the development of nanomedicines with enhanced therapeutic efficacy and reduced side effects in SLE therapy.