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1.
Orphanet J Rare Dis ; 15(1): 288, 2020 10 14.
Artigo em Inglês | MEDLINE | ID: mdl-33054853

RESUMO

BACKGROUND: Isolated macrodactyly is a severe congenital hand anomaly with functional and physiological impact. Known causative genes include PIK3CA, AKT1 and PTEN. The aim of this study is to gain insights into the genetics basis of isolated macrodactyly. RESULTS: We enrolled 24 patients with isolated macrodactyly. Four of them were diagnosed with Proteus syndrome based on skin presentations characteristic to this disease. Targeted next-generation sequencing was performed using patients' blood and affected tissues. Overall, 20 patients carry mosaic PIK3CA pathogenic variants, i.e. p.His1047Arg (N = 7), p.Glu542Lys (N = 6), p.Glu545Lys (N = 2), p.His1047Leu (N = 2), p.Glu453Lys (N = 1), p.Gln546Lys (N = 1) and p.His1047Tyr (N = 1). Four patients who met the diagnostic criteria of Proteus syndrome carry mosaic AKT1 p.Glu17Lys variant. Variant allele frequencies of these mosaic variants obtained through next-generation sequencing range from 10 to 33%. In genotype-phenotype correlation analysis of patients with PIK3CA variant, we found that patients with the macrodactyly of the lower limbs tend to carry PIK3CA variants located in the helical domain (P = 0.005). CONCLUSIONS: Mosaic PIK3CA and AKT1 variants can be found in all of our samples with isolated macrodactyly. Insights into phenotypic and genetic spectrum of isolated macrodactyly may be helpful in perusing a more precise and effective management of isolated macrodactyly.

2.
Orphanet J Rare Dis ; 15(1): 250, 2020 09 15.
Artigo em Inglês | MEDLINE | ID: mdl-32933559

RESUMO

BACKGROUND: We previously reported a novel clinically distinguishable subtype of congenital scoliosis (CS), namely, TBX6-associated congenital scoliosis (TACS). We further developed the TBX6-associated CS risk score (TACScore), a multivariate phenotype-based model to predict TACS according to the patient's clinical manifestations. In this study, we aimed to evaluate whether using the TACScore as a screening method prior to performing whole-exome sequencing (WES) is more cost-effective than using WES as the first-line genetic test for CS. METHODS: We retrospectively collected the molecular data of 416 CS patients in the Deciphering disorders Involving Scoliosis and COmorbidities (DISCO) study. A decision tree was constructed to estimate the cost and the diagnostic time required for the two alternative strategies (TACScore versus WES). Bootstrapping simulations and sensitivity analyses were performed to examine the distributions and robustness of the estimates. The economic evaluation considered both the health care payer and the personal budget perspectives. RESULTS: From the health care payer perspective, the strategy of using the TACScore as the primary screening method resulted in an average cost of $1074.2 (95%CI: $1044.8 to $1103.5) and an average diagnostic duration of 38.7d (95%CI: 37.8d to 39.6d) to obtain a molecular diagnosis for each patient. In contrast, the corresponding values were $1169.6 (95%CI: $1166.9 to $1172.2) and 41.4d (95%CI: 41.1d to 41.7d) taking WES as the first-line test (P < 0.001). From the personal budget perspective, patients who were predicted to be positive by the TACScore received a result with an average cost of $715.1 (95%CI: $594.5 to $835.7) and an average diagnostic duration of 30.4d (95%CI: 26.3d to 34.6d). Comparatively, the strategy of WES as the first-line test was estimated to have significantly longer diagnostic time with an average of 44.0d (95%CI: 43.2d to 44.9d), and more expensive with an average of $1193.4 (95%CI: $1185.5 to $1201.3) (P < 0.001). In 100% of the bootstrapping simulations, the TACScore strategy was significantly less costly and more time-saving than WES. The sensitivity analyses revealed that the TACScore strategy remained cost-effective even when the cost per WES decreased to $8.8. CONCLUSIONS: This retrospective study provides clinicians with economic evidence to integrate the TACScore into clinical practice. The TACScore can be considered a cost-effective tool when it serves as a screening test prior to performing WES.

3.
Mol Genet Genomic Med ; 8(10): e1453, 2020 10.
Artigo em Inglês | MEDLINE | ID: mdl-32815649

RESUMO

BACKGROUND: Congenital scoliosis (CS) is a spinal deformity due to vertebral malformations. Although insufficiency of TBX6 dosage contributes to a substantial proportion of CS, the molecular etiology for the majority of CS remains largely unknown. TBX6-mediated genes involved in the process of somitogenesis represent promising candidates. METHODS: Individuals affected with CS and without a positive genetic finding were referred to this study. Proband-only exome sequencing (ES) were performed on the recruited individuals, followed by analysis of TBX6-mediated candidate genes, namely MEOX1, MEOX2, MESP2, MYOD1, MYF5, RIPPLY1, and RIPPLY2. RESULTS: A total of 584 patients with CS of unknown molecular etiology were recruited. After ES analysis, protein-truncating variants in RIPPLY1 and MYF5 were identified from two individuals, respectively. In addition, we identified five deleterious missense variants (MYOD1, n = 4; RIPPLY2, n = 1) in TBX6-mediated genes. We observed a significant mutational burden of MYOD1 in CS (p = 0.032) compared with the in-house controls (n = 1854). Moreover, a potential oligogenic disease-causing mode was proposed based on the observed mutational co-existence of MYOD1/MEOX1 and MYOD1/RIPPLY1. CONCLUSION: Our study characterized the mutational spectrum of TBX6-mediated genes, prioritized core candidate genes/variants, and provided insight into a potential oligogenic disease-causing mode in CS.

4.
BMC Med Genet ; 21(1): 115, 2020 05 27.
Artigo em Inglês | MEDLINE | ID: mdl-32460719

RESUMO

BACKGROUND: Multiple epiphyseal dysplasia (MED) is a skeletal disorder characterized by delayed and irregular ossification of the epiphyses and early-onset osteoarthritis. At least 66% of the reported autosomal dominant MED (AD-MED) cases are caused by COMP mutations. METHODS: We recruited a four-generation Chinese family with early-onset hip osteoarthritis, flatfoot, brachydactyly, and mild short stature. An assessment of the family history, detailed physical examinations, and radiographic evaluations were performed on the proband and other family members, followed by the performance of whole-exome sequencing (WES). The pathogenicity of the candidate mutation was also analyzed. RESULTS: An AD-MED family with 10 affected members and 17 unaffected members was recruited. The main radiographic findings were symmetrical changes in the dysplastic acetabulum and femoral heads, irregular contours of the epiphyses, a shortened femoral neck, and flatfoot. Lower bone density was also observed in the ankle joints, wrist joints, and knees, as well as irregular vertebral end plates. In the proband, we identified the missense mutation c.1153G > T (p. Asp385Tyr), located in exon 11 of the COMP gene. This mutation was assessed as 'pathogenic' because of its low allele frequency and its high likelihood of co-segregation with disease in the reported family. Sanger sequencing validated the novel heterozygous mutation c.1153G > T (p. Asp385Tyr) in exon 11 of COMP in all affected individuals in the family. CONCLUSIONS: Our results underlined a key role of the Asp385 amino acid in the protein function of COMP and confirmed the pathogenicity of the COMP (c.1153G > T; p. Asp385Tyr) mutation in AD-MED disease. We have therefore expanded the known mutational spectrum of COMP and revealed new phenotypic information for AD-MED.


Assuntos
Proteína de Matriz Oligomérica de Cartilagem/genética , Família , Estudos de Associação Genética , Predisposição Genética para Doença , Mutação , Osteocondrodisplasias/diagnóstico , Osteocondrodisplasias/genética , Adolescente , Adulto , Idoso , Osso e Ossos/diagnóstico por imagem , Osso e Ossos/patologia , Proteína de Matriz Oligomérica de Cartilagem/química , Criança , Feminino , Estudos de Associação Genética/métodos , Humanos , Masculino , Pessoa de Meia-Idade , Modelos Moleculares , Linhagem , Fenótipo , Conformação Proteica , Relação Estrutura-Atividade , Sequenciamento Completo do Exoma , Adulto Jovem
5.
J Med Genet ; 2020 May 07.
Artigo em Inglês | MEDLINE | ID: mdl-32381727

RESUMO

BACKGROUND: Early-onset scoliosis (EOS), defined by an onset age of scoliosis less than 10 years, conveys significant health risk to affected children. Identification of the molecular aetiology underlying patients with EOS could provide valuable information for both clinical management and prenatal screening. METHODS: In this study, we consecutively recruited a cohort of 447 Chinese patients with operative EOS. We performed exome sequencing (ES) screening on these individuals and their available family members (totaling 670 subjects). Another cohort of 13 patients with idiopathic early-onset scoliosis (IEOS) from the USA who underwent ES was also recruited. RESULTS: After ES data processing and variant interpretation, we detected molecular diagnostic variants in 92 out of 447 (20.6%) Chinese patients with EOS, including 8 patients with molecular confirmation of their clinical diagnosis and 84 patients with molecular diagnoses of previously unrecognised diseases underlying scoliosis. One out of 13 patients with IEOS from the US cohort was molecularly diagnosed. The age at presentation, the number of organ systems involved and the Cobb angle were the three top features predictive of a molecular diagnosis. CONCLUSION: ES enabled the molecular diagnosis/classification of patients with EOS. Specific clinical features/feature pairs are able to indicate the likelihood of gaining a molecular diagnosis through ES.

6.
BMC Musculoskelet Disord ; 21(1): 220, 2020 Apr 11.
Artigo em Inglês | MEDLINE | ID: mdl-32278351

RESUMO

BACKGROUND: Klippel-Feil syndrome (KFS) represents a rare anomaly characterized by congenital fusion of the cervical vertebrae. The underlying molecular etiology remains largely unknown because of the genetic and phenotypic heterogeneity. METHODS: We consecutively recruited a Chinese cohort of 37 patients with KFS. The clinical manifestations and radiological assessments were analyzed and whole-exome sequencing (WES) was performed. Additionally, rare variants in KFS cases and controls were compared using genetic burden analysis. RESULTS: We primarily examined rare variants in five reported genes (GDF6, MEOX1, GDF3, MYO18B and RIPPLY2) associated with KFS and detected three variants of uncertain significance in MYO18B. Based on rare variant burden analysis of 96 candidate genes related to vertebral segmentation defects, we identified BAZ1B as having the highest probability of association with KFS, followed by FREM2, SUFU, VANGL1 and KMT2D. In addition, seven patients were proposed to show potential oligogenic inheritance involving more than one variants in candidate genes, the frequency of which was significantly higher than that in the in-house controls. CONCLUSIONS: Our study presents an exome-sequenced cohort and identifies five novel genes potentially associated with KFS, extending the spectrum of known mutations contributing to this syndrome. Furthermore, the genetic burden analysis provides further evidence for potential oligogenic inheritance of KFS.

7.
Med Sci Monit ; 26: e921611, 2020 Mar 16.
Artigo em Inglês | MEDLINE | ID: mdl-32218412

RESUMO

BACKGROUND Adolescent idiopathic scoliosis (AIS) is the most common spinal deformity, but its etiology is unclear. Multiple genetic mutations have been reported to be associated with AIS. MATERIAL AND METHODS We enrolled a cohort of 113 surgically treated AIS patients with available parental subjects from the Peking Union Medical College Hospital. We performed whole-exome sequencing in 10 trio families and whole-genome sequencing in 103 singleton patients. Luciferase assay was used to detect the functional alterations of candidate ESR1 and ESR2 variants. RESULTS Using a de novo strategy, a missense variant in ESR1 (c.868A>G) was selected as a candidate gene for AIS. The main Cobb angle of this patient was 41° (T6-T10). Another potential pathogenic variant in ESR2 (c.236T>C) was identified. The main curve of the patient was 45° at T10-L3. The transactivation capacities of the mutated ESR1 and ESR2 protein were both significantly decreased (p=0.026 and 0.014, respectively). CONCLUSIONS Potential pathogenic variants in ESR1 and ESR2 were identified in 113 AIS patients, suggesting that genetic mutations in ESR1/2 were associated with the risk of AIS.

8.
Hum Mutat ; 41(1): 182-195, 2020 01.
Artigo em Inglês | MEDLINE | ID: mdl-31471994

RESUMO

Congenital scoliosis (CS) is a birth defect with variable clinical and anatomical manifestations due to spinal malformation. The genetic etiology underlying about 10% of CS cases in the Chinese population is compound inheritance by which the gene dosage is reduced below that of haploinsufficiency. In this genetic model, the trait manifests as a result of the combined effect of a rare variant and common pathogenic variant allele at a locus. From exome sequencing (ES) data of 523 patients in Asia and two patients in Texas, we identified six TBX6 gene-disruptive variants from 11 unrelated CS patients via ES and in vitro functional testing. The in trans mild hypomorphic allele was identified in 10 of the 11 subjects; as anticipated these 10 shared a similar spinal deformity of hemivertebrae. The remaining case has a homozygous variant in TBX6 (c.418C>T) and presents a more severe spinal deformity phenotype. We found decreased transcriptional activity and abnormal cellular localization as the molecular mechanisms for TBX6 missense loss-of-function alleles. Expanding the mutational spectrum of TBX6 pathogenic alleles enabled an increased molecular diagnostic detection rate, provided further evidence for the gene dosage-dependent genetic model underlying CS, and refined clinical classification.

9.
Mol Genet Genomic Med ; 8(1): e1023, 2020 01.
Artigo em Inglês | MEDLINE | ID: mdl-31774634

RESUMO

BACKGROUND: The molecular and genetic mechanisms by which different single nucleotide variant alleles in specific genes, or at the same genetic locus, cause distinct disease phenotypes often remain unclear. Allelic truncating mutations of FBN1 could cause either classical Marfan syndrome (MFS) or a more complicated phenotype associated with Marfanoid-progeroid-lipodystrophy syndrome (MPLS). METHODS: We investigated a small cohort, encompassing two classical MFS and one MPLS subjects from China, whose clinical presentation included scoliosis potentially requiring surgical intervention. Targeted next generation sequencing was performed on all the participants. We analyzed the molecular diagnosis, clinical features, and the potential molecular mechanism involved in the MPLS subject in our cohort. RESULTS: We report a novel de novo FBN1 mutation for the first Chinese subject with MPLS, a more complicated fibrillinopathy, and two subjects with more classical MFS. We further predict that the MPLS truncating mutation, and others previously reported, is prone to escape the nonsense-mediated decay (NMD), while MFS mutations are predicted to be subjected to NMD. Also, the MPLS mutation occurs within the glucogenic hormone asprosin domain of FBN1. In vitro experiments showed that the single MPLS mutation p.Glu2759Cysfs*9 appears to perturb proper FBN1 protein aggregation as compared with the classical MFS mutation p.Tyr2596Thrfs*86. Both mutations appear to upregulate SMAD2 phosphorylation in vitro. CONCLUSION: We provide direct evidence that a dominant-negative interaction of FBN1 potentially explains the complex MPLS phenotypes through genetic and functional analysis. Our study expands the mutation spectrum of FBN1 and highlights the potential molecular mechanism for MPLS.

10.
J Hum Genet ; 65(3): 221-230, 2020 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-31827250

RESUMO

Congenital scoliosis (CS) is a form of scoliosis caused by congenital vertebral malformations. Genetic predisposition has been demonstrated in CS. We previously reported that TBX6 loss-of-function causes CS in a compound heterozygous model; however, this model can explain only 10% of CS. Many monogenic and polygenic CS genes remain to be elucidated. In this study, we analyzed exome sequencing (ES) data of 615 Chinese CS from the Deciphering Disorders Involving Scoliosis and COmorbidities (DISCO) project. Cosegregation studies for 103 familial CS identified a novel heterozygous nonsense variant, c.2649G>A (p.Trp883Ter) in FBN1. The association between FBN1 and CS was then analyzed by extracting FBN1 variants from ES data of 574 sporadic CS and 828 controls; 30 novel variants were identified and prioritized for further analyses. A mutational burden test showed that the deleterious FBN1 variants were significantly enriched in CS subjects (OR = 3.9, P = 0.03 by Fisher's exact test). One missense variant, c.2613A>C (p.Leu871Phe) was recurrent in two unrelated CS subjects, and in vitro functional experiments for the variant suggest that FBN1 may contribute to CS by upregulating the transforming growth factor beta (TGF-ß) signaling. Our study expanded the phenotypic spectrum of FBN1, and provided nove insights into the genetic etiology of CS.


Assuntos
Anormalidades Congênitas/genética , Fibrilina-1/genética , Predisposição Genética para Doença , Escoliose/genética , Criança , Pré-Escolar , Códon sem Sentido/genética , Anormalidades Congênitas/diagnóstico por imagem , Anormalidades Congênitas/fisiopatologia , Exoma/genética , Feminino , Estudos de Associação Genética , Humanos , Lactente , Masculino , Mutação , Mutação de Sentido Incorreto/genética , Linhagem , Escoliose/diagnóstico por imagem , Escoliose/fisiopatologia , Coluna Vertebral/diagnóstico por imagem , Coluna Vertebral/fisiopatologia , Fator de Crescimento Transformador beta/genética
11.
BMJ Open ; 9(10): e030726, 2019 10 28.
Artigo em Inglês | MEDLINE | ID: mdl-31662377

RESUMO

INTRODUCTION: Patient-reported outcome-based symptom monitoring and alerting have been attractive for patient care after a tumour-removal surgery. However, the implementation parameters of this patient-centred symptom management system in perioperative patients with lung cancer are still lacking. We aim to develop a perioperative symptom scale (PSS) for monitoring, to determine the optimal time points for symptom assessment and to define the alert thresholds for medical intervention. METHODS AND ANALYSIS: This study will prospectively recruit 300 patients undergoing lung cancer surgery in six hospitals. The MD Anderson Symptom Inventory-Lung Cancer Module (MDASI-LC) is used to collect longitudinal symptom data preoperatively, daily postoperatively during in-hospital stay and weekly after discharge until 4 weeks or the start of postoperative oncological therapy. Symptoms that change significantly over time will be generated as the PSS. We will determine the optimal time points for follow-up using the generalised linear mixed-effects models. The MDASI-LC interference-measured functional status will be used as the anchor for the alert thresholds. ETHICS AND DISSEMINATION: Ethics Committee of Sichuan Cancer Hospital approved this study on 16 October 2017 (No. SCCHEC-02-2017-042). The manuscript is based on the latest protocol of Version 3.0, 15 September 2019. The results of this study will be presented at medical conferences and published in peer-reviewed journals. TRIALS REGISTRATION NUMBER: NCT03341377.


Assuntos
Neoplasias Pulmonares/cirurgia , Medidas de Resultados Relatados pelo Paciente , Assistência Perioperatória/métodos , Complicações Pós-Operatórias/diagnóstico , Estudos de Coortes , Dispneia/diagnóstico , Dispneia/fisiopatologia , Dispneia/terapia , Fadiga/diagnóstico , Fadiga/fisiopatologia , Fadiga/terapia , Humanos , Neoplasias Pulmonares/fisiopatologia , Dor Pós-Operatória/diagnóstico , Dor Pós-Operatória/fisiopatologia , Dor Pós-Operatória/terapia , Complicações Pós-Operatórias/fisiopatologia , Complicações Pós-Operatórias/terapia , Estudos Prospectivos , Reprodutibilidade dos Testes , Avaliação de Sintomas
12.
Curr Gene Ther ; 19(4): 242-247, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31549955

RESUMO

OBJECTIVE: The genetic variations contributed to a substantial proportion of congenital vertebral malformations (CVM). SOX9 gene, a member of the SOX gene family, has been implicated in CVM. To study the SOX9 mutation in CVM patients is of great significance to explain the pathogenesis of scoliosis (the clinical manifestation of CVM) and to explore the pathogenesis of SOX9-related skeletal deformities. METHODS: A total of 50 singleton patients with CVM were included in this study. Exome Sequencing (ES) was performed on all the patients. The recurrent candidate variant of SOX9 gene was validated by Sanger sequencing. Luciferase assay was performed to investigate the functional changes of this variant. RESULTS: A recurrent rare heterozygous missense variant in SOX9 gene (NM_000346.3: c.1405A>G, p.M469V) which had not been reported previously was identified in three CVM patients who had the clinical findings of congenital scoliosis without deformities in other systems. This variant was absent from our in-house database and it was predicted to be deleterious (CADD = 24.5). The luciferase assay demonstrated that transactivation capacity of the mutated SOX9 protein was significantly lower than that of the wild-type for the two luciferase reporters (p = 0.0202, p = 0.0082, respectively). CONCLUSION: This SOX9 mutation (p.M469V) may contribute to CVM without other systematic deformity, which provides important implications and better understanding of phenotypic variability in SOX9-related skeletal deformities.


Assuntos
Anormalidades Congênitas/genética , Mutação de Sentido Incorreto , Fatores de Transcrição SOX9/genética , Doenças da Coluna Vertebral/genética , Coluna Vertebral/anormalidades , Adolescente , Criança , Pré-Escolar , China/epidemiologia , Anormalidades Congênitas/epidemiologia , Feminino , Heterozigoto , Humanos , Masculino , Prognóstico , Doenças da Coluna Vertebral/congênito , Doenças da Coluna Vertebral/epidemiologia , Coluna Vertebral/metabolismo , Coluna Vertebral/patologia
13.
BMJ Open ; 9(8): e030041, 2019 08 26.
Artigo em Inglês | MEDLINE | ID: mdl-31455710

RESUMO

INTRODUCTION: Surgery is one of the primary treatments for lung cancer. The postoperative symptom burden experienced by patients with lung cancer is substantial, seriously delaying their recovery from surgery and impairing their quality of life. Patient-reported outcome (PRO)-based symptom management is increasingly regarded as an optimal model for patient-centred care. Currently, clinical trial-based evidence involving early-phase (immediately after surgery for up to 1 month) symptom management of lung cancer is lacking. We propose a randomised trial to evaluate the effect of a PRO-based symptom-monitoring programme with overthreshold alerts and responses for postoperative recovery in patients with lung cancer. METHODS AND ANALYSIS: The study will recruit 160 patients with lung cancer from six hospitals. The patients will be randomly allocated to the intervention group or control group in a ratio of 1:1. Patients in the intervention group will receive PRO-based symptom management from the specialists when their reported target symptom (pain, coughing, fatigue, disturbed sleep and shortness of breath) scores reach the preset threshold (score ≥4). Patients in the control group will not generate alerts and will follow the standard procedures for symptom management. All patients will receive symptom assessments via the MD Anderson Symptom Inventory-lung cancer module on the day before surgery, daily after surgery and twice a week after discharge until 4 weeks or the start of postoperative oncological treatment. The primary outcome-mean symptom threshold events-will be compared between the intervention and control group via independent sample Student's t-test. ETHICS AND DISSEMINATION: The study was approved by the Ethics Committee of Sichuan Cancer Hospital on 22 November 2018 (No. SCCHEC-02-2018-045). This manuscript is based on V.2.0, 9 May 2019 of the protocol. The study results will be disseminated in publications in peer-reviewed journals and presentations at academic conferences. TRIALS REGISTRATION NUMBER: ChiCTR1900020846.


Assuntos
Neoplasias Pulmonares/cirurgia , Medidas de Resultados Relatados pelo Paciente , Qualidade de Vida , China , Gerenciamento Clínico , Humanos , Neoplasias Pulmonares/fisiopatologia , Neoplasias Pulmonares/psicologia , Estudos Multicêntricos como Assunto , Período Pós-Operatório , Ensaios Clínicos Controlados Aleatórios como Assunto , Índice de Gravidade de Doença , Carga Tumoral
14.
Cell Death Dis ; 9(10): 964, 2018 09 20.
Artigo em Inglês | MEDLINE | ID: mdl-30237478

RESUMO

Disruption of the nursery function in Sertoli cells (SCs) by reducing lactate production, a preferred energy substrate for developed germ cells (spermatocytes and spermatids), is tightly associated with spermatogenic failure such as SC-only syndrome (SCOS). However, whether this complicated pathogenesis is regulated by certain miRNAs at the post-transcriptional level remain fascinating but largely unknown. Here we show for the first time that mmu-miR-320-3p was exclusively expressed in murine SCs and this expression was significantly induced in busulphan-treated murine testis. The most efficient stimulatory germ cell types for the induction of apoptosis-elicited mmu-miR-320-3p expression were meiotic spermatocytes and haploid spermatids. Functionally, forced expression of the exogenous mmu-miR-320-3p in SCs compromises male fertility by causing oligozoospermia and defection of sperm mobility. Mechanistically, mmu-miR-320-3p negatively regulates lactate production of SCs by directly inhibiting glucose transporter 3 (GLUT3) expression. Thus, dysregulation of mmu-miR-320-3p/GLUT3 cascade and consequently of lactate deficiency may be a key molecular event contributing the germ cell loss by SC dysfunction. Future endeavor in the continuous investigation of this important circulating miRNA may shed novel insights into epigenetic regulation of SCs nursery function and the etiology of azoospermia, and offers novel therapeutic and prognostic targets for SCOS.


Assuntos
Transportador de Glucose Tipo 3/metabolismo , MicroRNAs/metabolismo , Células de Sertoli/metabolismo , Animais , Epigênese Genética/genética , Epigênese Genética/fisiologia , Perfilação da Expressão Gênica , Transportador de Glucose Tipo 3/genética , Lactatos/metabolismo , Masculino , Camundongos , MicroRNAs/genética , Transdução de Sinais/genética , Transdução de Sinais/fisiologia
15.
Life Sci ; 207: 355-363, 2018 Aug 15.
Artigo em Inglês | MEDLINE | ID: mdl-29940243

RESUMO

AIMS: The pleckstrin homology domain leucine-rich repeat protein phosphatase 1 (PHLPP1) specifically regulates phospho-Ser473 of protein kinase B (PKB, Akt) opposing cell survival during myocardial ischemia/reperfusion (I/R). Previous studies demonstrated PHLPP1 expression level was controlled by several mechanisms. However, the regulation mechanism of cardiac PHLPP1 expression following myocardial I/R remains unknown. MAIN METHODS: The current study utilized the mouse model of myocardial I/R injury in vivo and the neonatal rat ventricular myocytes (NRVMs) of hypoxia/reoxygenation (H/R) injury in vitro. Expression of PHLPP1, nuclear factor-kappa B (NF-κB) and pNF-κB were determined by western blot. The expression of PHLPP1 and translocation of NF-κB was assessed by immunofluorescence. Chromatin immunoprecipitation (ChIP) assay was used to detect the binding of NF-κB to the promoter region of phlpp1 gene. KEY FINDINGS: Myocardial I/R had no effect on cardiac PHLPP1 expression following I/R (30 min/2 h) but decreased after 4 h reperfusion. In vitro, H/R (4 h/1 h) and tumor necrosis factor-alpha (TNF-α)-stimulation resulted in upregulation of PHLPP1 in NRVMs, which was blocked with etanercept. Yet, H2O2-induced oxidative stress had no obvious effect on PHLPP1 expression of NRVMs at early stage but N-acetylcysteine (NAC) pretreatment increased PHLPP1 levels after 4 h H2O2 stimulation. TNF-α and H/R led to both expression and transcriptional activity of NF-κB, accompany with higher expression of PHLPP1. Pyrrolidine dithiocarbamate (PDTC), a NF-κB inhibitor, prevented the response not only in TNF-α-treated cardiomyocytes but also in H/R-treated group. SIGNIFICANCE: These results implicated that TNF-α involved in cardiac PHLPP1 upregulation during reoxygenation, which was mediated by NF-κB transcriptional activity.


Assuntos
Isquemia Miocárdica/metabolismo , Subunidade p50 de NF-kappa B/metabolismo , NF-kappa B/metabolismo , Proteínas Nucleares/metabolismo , Fosfoproteínas Fosfatases/metabolismo , Traumatismo por Reperfusão/metabolismo , Fator de Necrose Tumoral alfa/metabolismo , Animais , Animais Recém-Nascidos , Modelos Animais de Doenças , Regulação da Expressão Gênica , Ventrículos do Coração/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Células Musculares/citologia , Células Musculares/metabolismo , Regiões Promotoras Genéticas , Proteínas Proto-Oncogênicas c-akt/metabolismo , Ratos , Ratos Sprague-Dawley , Serina/química
16.
Oncotarget ; 8(26): 43023-43034, 2017 Jun 27.
Artigo em Inglês | MEDLINE | ID: mdl-28498809

RESUMO

OBJECTIVE: Follistatin-like protein 1 (FSTL1) is a well-known mediator of inflammation. Intervertebral disc disease is an inflammatory disorder. Here, we investigated the role of FSTL1 in the intervertebral discs inflammation. METHODS: Expression of FSTL1 in nucleus pulposus tissues from rats and human was determined by immunohistochemistry staining and western blot analysis. The expression levels of tumor necrosis factor-α (TNF-α), interleukin1-ß (IL-1ß) and matrix metalloproteinase 13 (MMP-13) in human and rat nucleus pulposus tissues were measured by immunohistochemistry staining. The mitogen-activated protein kinase (MAPK) and nuclear factor-kappa B (NFκB) signaling pathways were detected by western blotting. RESULTS: FSTL1 serum levels were significantly increased in lumbar disc herniation patients and had a positive correlation with Visual Analogue Scores. Additionally, FSTL1 expression was significantly increased in extrusion group compared with protrusion and control groups. Furthermore, FSTL1 expression was significantly increased in intervertebral disc degeneration models of rats. Immunohistochemistry staining demonstrated that the levels of TNF-α, IL-1ß and MMP-13 were increased in the pathogenesis of intervertebral disc disease. Recombinant human FSTL1 significantly increased the production of proinflammatory cytokines in vitro. In addition, FSTL1 promoted inflammation by activating c-Jun N-terminal kinase (JNK), extracellular regulated protein kinases 1/2(ERK1/2) and NFκB signaling. CONCLUSIONS: These data imply that FSTL1 expression was increased in the pathogenesis of intervertebral disc disease. Importantly, FSTL1 promoted inflammatory catabolism in the nucleus pulposus by activating JNK, ERK 1/2/MAPK and NFκB signaling.


Assuntos
Proteínas Relacionadas à Folistatina/metabolismo , Inflamação/etiologia , Inflamação/metabolismo , Proteínas Quinases Ativadas por Mitógeno/metabolismo , NF-kappa B/metabolismo , Núcleo Pulposo/citologia , Transdução de Sinais , Animais , Biomarcadores , Citocinas/sangue , Citocinas/metabolismo , Feminino , Proteínas Relacionadas à Folistatina/sangue , Proteínas Relacionadas à Folistatina/genética , Expressão Gênica , Humanos , Inflamação/patologia , Mediadores da Inflamação/sangue , Mediadores da Inflamação/metabolismo , Degeneração do Disco Intervertebral/etiologia , Degeneração do Disco Intervertebral/metabolismo , Degeneração do Disco Intervertebral/patologia , Deslocamento do Disco Intervertebral/etiologia , Deslocamento do Disco Intervertebral/metabolismo , Deslocamento do Disco Intervertebral/patologia , Masculino , Ratos , Transdução de Sinais/efeitos dos fármacos , Fator de Necrose Tumoral alfa/metabolismo , Fator de Necrose Tumoral alfa/farmacologia
17.
Inflammation ; 40(2): 612-622, 2017 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-28233079

RESUMO

This study was conducted in order to investigate the function of IL-21 in intervertebral disc degeneration. The serum concentration of IL-21 in patients with lumbar disc herniation (LDH) was examined by ELISA. Immunohistochemistry and western blot analysis were performed to detect the expression of IL-21, a disintegrin and metalloproteinase with thrombospondin motifs (ADAMTS-7), and tumor necrosis factor alpha (TNF-α) in degenerated intervertebral disc (IVD) tissues of human and rat. Moreover, nucleus pulposus (NP) cells were treated with 0, 10, 100, and 1000 ng/mL of IL-21 cytokine with and without AG490. TNF-α, ADAMTS-7, and matrix metalloproteinases-13 (MMP-13) mRNA expression was determined by RT-PCR. The expression of signal transducers and activators of transcription, STAT-1, STAT-3, and STAT-5b, was detected by western blot. IL-21 concentration level is higher in the degenerated group and positively correlates with the visual analog score (VAS). IL-21, ADAMTS-7, and TNF-α can be detected in the degenerative NP tissues in both human and rat degenerated NP tissues. The mRNA expression of ADAMTS-7, TNF-α, and MMP-13 was enhanced after stimulation with IL-21. Compared to control, STAT-1, STAT-3, and STAT-5b expression was also enhanced after IL-21 treatment, with STAT-3 being the most significantly enhanced; furthermore, expression was significantly reduced after treatment with AG490. The mRNA expression of TNF-α was markedly reduced after treatment with AG490 compared to treatment with IL-21 only. IL-21 is involved in the pathological development of IVD degeneration and IL-21 could aggravate IVD degeneration by stimulating TNF-α through the STAT signaling pathway.


Assuntos
Interleucinas/fisiologia , Degeneração do Disco Intervertebral/etiologia , Janus Quinases/metabolismo , Fatores de Transcrição STAT/metabolismo , Transdução de Sinais , Fator de Necrose Tumoral alfa/metabolismo , Adulto , Animais , Células Cultivadas , Relação Dose-Resposta a Droga , Feminino , Humanos , Interleucinas/administração & dosagem , Interleucinas/sangue , Degeneração do Disco Intervertebral/metabolismo , Masculino , Pessoa de Meia-Idade , Núcleo Pulposo/efeitos dos fármacos , Núcleo Pulposo/patologia , RNA Mensageiro/análise , Ratos , Fatores de Transcrição STAT/análise , Fator de Necrose Tumoral alfa/genética , Tirfostinas/farmacologia
18.
Spine (Phila Pa 1976) ; 41(21): 1631-1640, 2016 Nov 01.
Artigo em Inglês | MEDLINE | ID: mdl-27802252

RESUMO

STUDY DESIGN: Based on human disc surgical samples and isolated cells in vitro, we undertook a descriptive and mechanistic investigation of proinflammatory effects of interleukin (IL)-9 in intervertebral disc (IVD) degeneration. OBJECTIVE: To investigate the proinflammatory role of IL-9 in the pathological process of IVD degeneration. SUMMARY OF BACKGROUND DATA: IL-9 is known as a pleiotropic cytokine that regulates the human pathogenesis of inflammatory and autoimmune diseases. However, whether IL-9 cytokine is involved in the immuno-inflammatory pathogenesis of IVD degeneration is unclear. METHODS: The IVD samples were obtained from 45 patients. Immunohistochemistry, western blot, and real-time Polymerase Chain Reaction (PCR) were performed to detect the expression of IL-9 and tumor necrosis factor alpha (TNF-α) in the degenerated IVDs. Moreover, nucleus pulposus (NP) cells were treated with 0, 1, 10, and 100 ng/mL IL-9 cytokine and stimulated with IL-9 alone at 100 ng/mL for 0, 12, 24, and 48 hours. TNF-α expression was determined by immunofluorescence staining, western blot, and real-time PCR, respectively. The amounts of TNF-α and prostaglandin E2 (PGE2) in the supernatant were quantified by enzyme-linked immunosorbent assay. Additionally, Spearman correlation analyses were performed to analyze the correlation between Pfirrmann grading score of the involved degenerated IVDs and serum levels of IL-9. RESULTS: The expressions of IL-9 and TNF-α in degenerated IVD tissues were dramatically elevated in comparison with the control. IL-9 significantly up-regulated the TNF-α and PGE2 secretion of NP cells in dose- and time-dependent manner. Moreover, there is a positive correlation between IL-9 serum level and severity of involved IVD degeneration. CONCLUSION: Our findings suggest that IL-9 may play a potential role in the inflammatory processes of IVD degeneration. IL-9 may be involved in the IVD degeneration, at least in part, though stimulating the release of TNF-α and PGE2 in NP cells. LEVEL OF EVIDENCE: N/A.


Assuntos
Dinoprostona/metabolismo , Interleucina-9/farmacologia , Degeneração do Disco Intervertebral/metabolismo , Disco Intervertebral/efeitos dos fármacos , Fator de Necrose Tumoral alfa/metabolismo , Adulto , Feminino , Humanos , Inflamação/metabolismo , Interleucina-9/metabolismo , Disco Intervertebral/metabolismo , Masculino , Pessoa de Meia-Idade , Adulto Jovem
19.
Inflammation ; 39(6): 1997-2007, 2016 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-27590238

RESUMO

Matrix metalloproteinase 13 (MMP-13) plays an important role in the process of pro-inflammatory cytokine-induced intervertebral disc degeneration (IDD). This study examined the effect of IL-17 on the regulation of MMP-13 and the extracellular matrix (ECM) in the intervertebral disc (IVD). We then examined whether salubrinal, a known inhibitor of eIF2α dephosphorylation, inhibited the IL-17-induced changes mentioned above. Furthermore, we demonstrated a potential therapeutic role for salubrinal in alleviating the chronic inflammatory-dependent degenerative state commonly observed in IDD. After inflammatory distress with IL-17, RT-PCR and western blot were employed to investigate the expression of MMP-13, collagen type II (COL2A1), collagen type I (COL1A1), and aggrecan (ACAN) in nucleus pulpous (NP) tissue. Activation of the NF-kB pathway was measured by western blot and immunocytochemistry following IL-17 treatment. We also examine the level of eIF2α phosphorylation after IL-17 treatment with or without salubrinal. Then, we investigated interactions of the NF-kB pathway to eIF2α phosphorylation. Moreover, we employed salubrinal and a specific inhibitor of NF-kB (BAY11-7082) to evaluate their effects on IL-17-driven regulation of MMP-13 and the ECM, as well as on the activation of NF-kB. The results showed that IL-17 increased the production of MMP-13 and decreased expression of COL2A1 and ACAN via the NF-kB pathway. Either IL-17 or salubrinal increased the level of eIF2α phosphorylation, but the effects of BAY11-7082 on the level of p-eIF2α were not detectable. BAY11-7082 and salubrinal significantly suppressed IL-17-driven intervertebral disc degeneration. Furthermore, salubrinal produced stronger effects than BAY11-7082. These results imply the potential involvement of IL-17 in IDD through activation of NF-kB signaling, which successively upregulated the expression of MMP-13 and led to the degradation of the ECM. Furthermore, salubrinal can inhibit this process through inhibition of NF-kB activation that is not directly linked to eIF2α phosphorylation, suggesting a potential therapeutic role in IDD.


Assuntos
Cinamatos/farmacologia , Matriz Extracelular/metabolismo , Interleucina-17/farmacologia , Metaloproteinase 13 da Matriz/metabolismo , NF-kappa B/metabolismo , Núcleo Pulposo/citologia , Transdução de Sinais/efeitos dos fármacos , Tioureia/análogos & derivados , Regulação para Cima/efeitos dos fármacos , Agrecanas/análise , Cinamatos/uso terapêutico , Colágeno Tipo I/análise , Colágeno Tipo II/análise , Fator de Iniciação 2 em Eucariotos/metabolismo , Humanos , Degeneração do Disco Intervertebral/tratamento farmacológico , Metaloproteinase 13 da Matriz/efeitos dos fármacos , Nitrilos/farmacologia , Núcleo Pulposo/efeitos dos fármacos , Núcleo Pulposo/metabolismo , Sulfonas/farmacologia , Tioureia/farmacologia , Tioureia/uso terapêutico
20.
J Transl Med ; 14: 77, 2016 Mar 17.
Artigo em Inglês | MEDLINE | ID: mdl-26988982

RESUMO

BACKGROUND: Low back pain and sciatica caused by intervertebral disc (IVD) disease are associated with inflammatory responses. The cytokine interleukin 17 (IL-17) is elevated in herniated and degenerated IVD tissues and acts as a regulator of disc inflammation. The objective of this study was to investigate the involvement of IL-17A in IVD inflammatory response and to explore the mechanisms underlying this response. METHODS: Cells were isolated from nucleus pulposus (NP) tissues collected from patients undergoing surgeries for IVD degeneration. The concentrations of COX2 and PGE2, as well as of select proteins involved in the mitogen-activated protein kinase (MAPK)/activating protein-1 (AP-1) pathway, were quantified in NP cells after exposure to IL-17 with or without pretreatment with MAPK or AP-1 inhibitors. RESULTS: Our results showed that IL-17A increased COX2 expression and PGE2 production via the activation of MAPKs, including p38 kinase and Jun N-terminal kinase (JNK). Moreover, IL-17A-induced COX2 and PGE2 production was shown to rely on p38/c-Fos and JNK/c-Jun activation in an AP-1-dependent manner. CONCLUSION: In summary, our results indicate that IL-17A enhances COX2 expression and PGE2 production via the p38/c-Fos and JNK/c-Jun signalling pathways in NP cells to mediate IVD inflammation.


Assuntos
Inflamação/patologia , Interleucina-17/farmacologia , Disco Intervertebral/patologia , Proteínas Quinases JNK Ativadas por Mitógeno/metabolismo , Proteínas Proto-Oncogênicas c-fos/metabolismo , Fator de Transcrição AP-1/metabolismo , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismo , Adulto , Células Cultivadas , Ciclo-Oxigenase 2/metabolismo , Dinoprostona/biossíntese , Ativação Enzimática/efeitos dos fármacos , Feminino , Humanos , Inflamação/metabolismo , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Masculino , Pessoa de Meia-Idade , Modelos Biológicos , Fosforilação/efeitos dos fármacos , Fatores de Tempo , Regulação para Cima/efeitos dos fármacos , Adulto Jovem
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