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1.
Hum Mutat ; 40(11): 2001-2006, 2019 11.
Artigo em Inglês | MEDLINE | ID: mdl-31292994

RESUMO

Empty follicle syndrome (EFS) is a condition in which no oocyte is retrieved from mature follicles after proper ovarian stimulation in an in vitro fertilization procedure. Genetic evidence accumulates for the etiology of recurrent EFS without pharmacological or iatrogenic problems. In this study, we present two infertile sisters in a family with EFS after three cycles of standard ovarian stimulation with human chorionic gonadotrophin and/or gonadotropin-releasing hormone agonist therapy. Via whole-exome sequencing and cosegregation test, we identified compound heterozygous mutations in the gene of ZP1 in both of the infertile sisters. Coimmunoprecipitation tests and homology modeling analysis confirmed that both mutated ZP1 disrupt the formation of oocyte zona pellucida by interrupting the interaction among ZP1, ZP2, and ZP3. We thus propose that the specific mutations in ZP1 gene render a causality for the intractable EFS.

2.
Zhongguo Shi Yan Xue Ye Xue Za Zhi ; 27(3): 904-910, 2019 Jun.
Artigo em Chinês | MEDLINE | ID: mdl-31204952

RESUMO

OBJECTIVE: To investigate the gene mutations types and the clinical characteristics in 3 patients with hereditary coagulation factor Ⅶ deficiency. METHODS: The phenotype diagnosis was validated by detecting the coagulation parameters including prothrombin time (PT),activated partial thromboplastin time (APTT), fibrinogen (FIB), FⅦ activity (FⅦ: C) and specific antigens (FⅦ: Ag) of proband and its family members. All exons, exon-intron boundaries, 5' untranslated regions and 3' untranslated regions of F7 gene were amplified with PCR. Potential mutations were detected by direct sequencing of purified PCR products. Suspected mutations were confirmed by sequencing of the opposite strand. RESULTS: A total of 5 different mutations were identified in 3 patients with hereditary coagulation factor Ⅶ deficiency and family members, including 4 misssense mutations and 1 splice site mutation. Out of 3 cases of hereditary coagulation factor Ⅶ deficiency 2 had double heterozygous mutation, I had homozygous mutations. Patient 1 had p.His408Gln with p.Arg413Gln double heterozygous mutations, her sister had p.His408Gln with p.Arg413Gln double heterozygous mutations, another one had p.His408Gln mono-heterozygous mutation, their correspo FⅦ: C were 5%, 3%, 75%. Patient 2 had p.Arg364Gln with p.His408Gln double heterozygous mutations, her brother had p.Arg364Gln with IVS6-1G>A double heterozygous mutations, their corresponding FⅦ: C were 2.0%, 2.0%. Patient 3 had p.Arg337Cys homozygous mutation, FⅦ: C was 3.0%. CONCLUSION: A total of 5 different mutations were identified in 3 patients with hereditary coagulation factor Ⅶ deficiency, the p.His408Gln is a common mutation, the FⅦ: C and FⅦ: Ag have no correlation with clinical phenotypes.


Assuntos
Deficiência do Fator VII , Fator VII , Feminino , Heterozigoto , Homozigoto , Humanos , Masculino , Mutação , Linhagem , Fenótipo
3.
Epigenetics ; 14(8): 791-803, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31088315

RESUMO

Renal cell carcinoma (RCC) is a common malignant tumour affecting the urinary system, and multidrug resistance is one of the major reasons why chemotherapy for this type of cancer often fails. Previous studies have shown that loss of the human organic cation transporter OCT2 is the main factor contributing to oxaliplatin resistance in RCC, and that DNA hypermethylation and histone methylation play important roles in the transcriptional repression of OCT2 in RCC. In this study, we found that histone acetylation also regulates OCT2 repression in RCC and elucidated the underlying mechanisms. In normal renal cells, HDAC7 combines with MYC at the OCT2 promoter, resulting in a decrease in free HDAC7, which in turn increases the levels of H3K18ac and H3K27ac at the OCT2 promotor and activates OCT2 expression. In RCC cells, however, the interaction between HDAC7 and MYC does not occur, which leads a high abundance of HDAC7 and low levels of H3K18ac and H3K27ac at the OCT2 promoter, thereby resulting in the inhibition of OCT2 transcription. We found that combined treatment using the DNA methylation inhibitor decitabine and the histone deacetylase inhibitor vorinostat significantly increased the expression of OCT2 in RCC cell lines, which sensitized these cells to oxaliplatin. We accordingly propose that the combination of anticancer agents and epigenetic drugs can provide a novel chemotherapeutic regimen.

4.
RNA Biol ; 16(7): 940-949, 2019 07.
Artigo em Inglês | MEDLINE | ID: mdl-30951404

RESUMO

SLC47A2 encodes MATE 2-K in the kidney, which mediates the secretion of certain endogenous and exogenous compounds. SLC47A2 was dramatically repressed in patients with renal cell carcinoma (RCC), and a lower level of SLC47A2 might act as a negative prognostic marker, although the mechanism is not well understood. In this study, we aimed to investigate the mechanism via which SLC47A2 is downregulated in RCC. Based on the annotation information of the SLC47A2 locus available in the UCSC genome browser database, we identified a novel lncRNA, which is transcribed from the SLC47A2 locus and named it SANT1. Overexpression and knock-down assays were performed to investigate the effects of SANT1 on cis-regulation of SLC47A2. We verified the direct binding between SANT1 and SFPQ/E2F1/HDAC1 using the cross-linking and immunoprecipitation (CLIP) assay. Chromatin immunoprecipitation was performed to confirm the molecular mechanism via which SANT1 activates the transcription of the SLC47A2 coding region. We observed that SANT1 can cis-regulate its own genetic locus. In tumour-adjacent tissues, the SLC47A2 locus highly expresses SANT1, which can remove the regulatory SFPQ/E2F1/HDAC1 suppressor complex from the promoter region, thereby significantly increasing the levels of the H3K27ac modification and RNAPII binding. Owing to a low SANT1 level, the binding of this inhibitory complex in the promoter region is upregulated in RCC, which results in silencing of the SLC47A2 coding region. In conclusion, we identified a novel lncRNA and elucidated the mechanism via which it regulates SLC47A2 expression in RCC.


Assuntos
Carcinoma de Células Renais/genética , Fator de Transcrição E2F1/metabolismo , Regulação Neoplásica da Expressão Gênica , Histona Desacetilase 1/metabolismo , Proteínas de Transporte de Cátions Orgânicos/metabolismo , Fator de Processamento Associado a PTB/metabolismo , Regiões Promotoras Genéticas , RNA Longo não Codificante/metabolismo , Sequência de Bases , Linhagem Celular Tumoral , Células HEK293 , Humanos , Neoplasias Renais/genética , Modelos Biológicos , Conformação de Ácido Nucleico , Proteínas de Transporte de Cátions Orgânicos/genética , Ligação Proteica , RNA Longo não Codificante/química , RNA Longo não Codificante/genética , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Regulação para Cima/genética
5.
Zhonghua Yi Xue Yi Chuan Xue Za Zhi ; 36(3): 221-224, 2019 Mar 10.
Artigo em Chinês | MEDLINE | ID: mdl-30835350

RESUMO

OBJECTIVE: To explore molecular etiology and clinical characteristics of two pedigrees affected with hereditary factor VII(FVII) deficiency. METHODS: The nine exons and flanking sequences of the F7 gene of the probands were amplified by PCR. The amplicons were analyzed by direct sequencing. Suspected mutations were subjected to SWISS-MODEL modeling and analysis of protein structure change by Pymol software and conservation of amino acids across various species. RESULTS: For proband of pedigree 1, the prothrombin time (PT), FVII activity (FVII:C) and FVII antigen (FVII:Ag) were 36.3 s, 3%, 53.56%, respectively. Sequencing revealed a compound heterozygous variants of c.80_81delCT and c.1371G>T(p.Arg439Ser). His son carried a heterozygous c.1371G>T (p.Arg439Ser) variant. For proband of pedigree 2, the PT, FVII:C and FVII:Ag were 22.3 s, 4%, 1.58%, respectively. Sequencing has revealed a compound heterozygous c.278G>T(p.Arg75Met) missense variant in exon 3 and c.1278T>G (p.His408Gln) in exon 9 of the F7 gene. His mother and son both carried a heterozygous c.278G>T(p.Arg75Met) variant. Three-dimensional simulation and homology analysis revealed that the p.Arg439Ser and p.Arg75Met can respectively alter part of hydrogen bonds and two highly conserved amino acids. CONCLUSION: Two novel heterozygous missense variants of the F7 gene [c.1371G>T(p.Arg439Ser) and c.278G>T(p.Arg75Met)] probably account for the decrease of factor VII in the two pedigrees.


Assuntos
Deficiência do Fator VII , Grupo com Ancestrais do Continente Asiático , Fator VII , Genótipo , Heterozigoto , Humanos , Mutação , Linhagem
7.
Oncol Lett ; 14(6): 7417-7424, 2017 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-29344182

RESUMO

Multiple studies have reported the prognostic association of certain inflammatory factors with various types of cancer. The present study assessed the prognostic value of the C-reactive protein (CRP)/albumin (Alb) ratio and the neutrophil/lymphocyte ratio (NLR), separately and in combination, in gastric cancer (GC). A total of 337 cases pathologically diagnosed with gastric adenocarcinoma were retrospectively evaluated. The clinicopathological and prognostic relevance of the CRP/Alb ratio and NLR and their combination were analyzed. The optimal cut-off values of the CRP/Alb ratio and NLR were 0.38 and 3.14, respectively. High CRP/Alb ratio (≥0.38) and NLR (≥3.14) values were associated with increased tumor invasion, more distant metastasis and a more advanced tumor-node-metastasis stage (all P<0.05). In addition, a high NLR value was also associated with increased tumor size (P=0.02). The CRP/Alb ratio (≥0.38/<0.38) and NLR (≥3.14/<3.14) were independent prognostic factors for overall survival time (OS) in GC by multivariate analysis (P=0.005 and P=0.001). Using the CRP/Alb ratio and NLR classification, patients were stratified into three subgroups with different OS time (P<0.001), which were identified as independent prognostic variables in multivariate analysis (P<0.001). The present study demonstrated that the CRP/Alb ratio and NLR were independent prognostic factors for OS in patients with GC. The combination of these indexes was associated with significant prognostic value and may further stratify prognosis.

8.
Drug Metab Dispos ; 45(1): 109-117, 2017 01.
Artigo em Inglês | MEDLINE | ID: mdl-27821436

RESUMO

In recent years, finding effective biomarkers for identifying early stage cancer and predicating prognosis is crucial for renal cell carcinoma (RCC) diagnosis and treatment. In this study, a dramatic decrease of the solute carrier family 47 member 2 (SLC47A2) mRNA in RCC comparing with the paired adjacent nontumor tissues from patients at low Tumor Node Metastasis stage was observed. Thus, patients with SLC47A2 transcriptional repression are susceptible to RCC. Little is known about the regulation mechanism of SLC47A2 We found that it was a bivalent gene that was enriched with both histone H3 lysine 4 trimethylation (H3K4me3) and lysine 27 trimethylation (H3K27me3). Loss of mixed lineage leukemia 1 binding at the gene promoter caused decreased H3K4me3 enrichment and H3K4me3/H3K27me3 ratio, and subsequently repressed the expression of SLC47A2 These two epigenetic markers modulated the expression of SLC47A2 simultaneously, suggesting the regulation pattern for bivalent genes. Histone H3 lysine 27 acetylation also contributed to the expression of SLC47A2 An E2F1-histone deacetylase 10 complex catalyzed deacetylation of H3K27, then prevented the enrichment of H3K4me3, and finally reduced SLC47A2 expression. Consequently, the combined effect of all these factors determined SLC47A2 transcriptional repression in RCC tissues.


Assuntos
Carcinoma de Células Renais/metabolismo , Histonas/metabolismo , Neoplasias Renais/metabolismo , Lisina/metabolismo , Proteínas de Transporte de Cátions Orgânicos/genética , Transcrição Genética , Acetilação , Carcinoma de Células Renais/genética , Linhagem Celular Tumoral , Imunoprecipitação da Cromatina , Epigênese Genética , Células HEK293 , Histonas/genética , Humanos , Neoplasias Renais/genética , Metilação
9.
Sci Transl Med ; 8(348): 348ra97, 2016 07 20.
Artigo em Inglês | MEDLINE | ID: mdl-27440728

RESUMO

Renal cell carcinoma (RCC) is known for its multidrug resistance. Using data obtained from the cancer transcriptome database Oncomine and the proteome database The Human Protein Atlas, we identified the repression of organic cation transporter OCT2 as a potential factor contributing to oxaliplatin resistance in RCC. By analyzing OCT2 expression in collected patient tissues and commercial tissue microarray specimens, we demonstrated OCT2 repression in RCC at both transcription and protein levels. Epigenetic analysis revealed that the repressed OCT2 promoter in RCC is characterized by hypermethylated CpG islands and the absence of H3K4 methylation. Further mechanistic studies showed that DNA hypermethylation blocked MYC activation of OCT2 by disrupting its interaction with the E-Box motif, which prevented MYC from recruiting MLL1 to catalyze H3K4me3 at the OCT2 promoter and resulted in repressed OCT2 transcription. Targeting this mechanism, we designed a sequential combination therapy and demonstrated that epigenetic activation of OCT2 by decitabine sensitizes RCC cells to oxaliplatin both in vitro and in xenografts. Our study highlights the potential of translating "omics" data into the development of targeted therapies.


Assuntos
Carcinoma de Células Renais/genética , Metilação de DNA/genética , Transportador 2 de Cátion Orgânico/metabolismo , Compostos Organoplatínicos/farmacologia , Ilhas de CpG/efeitos dos fármacos , Ilhas de CpG/genética , Metilação de DNA/efeitos dos fármacos , Epigenômica/métodos , Humanos , Imuno-Histoquímica , Técnicas In Vitro , Transportador 2 de Cátion Orgânico/genética , Oxaliplatina , Regiões Promotoras Genéticas/efeitos dos fármacos , Regiões Promotoras Genéticas/genética , Análise Serial de Tecidos
10.
Zhejiang Da Xue Xue Bao Yi Xue Ban ; 43(2): 160-3, 2014 Mar.
Artigo em Chinês | MEDLINE | ID: mdl-24782371

RESUMO

OBJECTIVE: To determine the enantiomeric impurity contents of domestic timolol maleate in bulk drugs and eye drops. METHODS: Enantiomer impurity of timolol was assayed by chiral high performance liquid chromatography. The chromatographic conditions were as follows:chiralcel OD chiral column (4.6 mm ×150 mm, 5µm), detection wavelength:297 nm, mobile phase:hexane-isopropanol-diethylamine (480:20:1), column temperature:25 ℃, flow rate:1.0 ml/min, sample injection volume:5 µl. RESULTS: The resolution between R- and S-timolol was more than 4. The enantiomeric impurity contents were less than 0.67% on average in two batches of timolol maleate bulk drugs, and 0.31% on average in three batches of timolol maleate eye drops. CONCLUSION: Enantiomeric impurity contents in each batch of products all meet European Pharmacopoeia criteria, which can be used as references in Chinese Pharmacopoeia criteria.


Assuntos
Cromatografia Líquida de Alta Pressão/métodos , Contaminação de Medicamentos , Soluções Oftálmicas/análise , Timolol/análise , Soluções Oftálmicas/normas , Estereoisomerismo , Timolol/normas
11.
Zhejiang Da Xue Xue Bao Yi Xue Ban ; 43(2): 164-7, 2014 Mar.
Artigo em Chinês | MEDLINE | ID: mdl-24782372

RESUMO

OBJECTIVE: To determine the contents of L-enantiomer impurity in valaciclovir hydrochloride. METHODS: Valaciclovir enantiomers were separated and determined by using chiral high performance liquid chromatography. Chromatographic conditions were as follows:CROWNPAK(®) CR(+) chiral column (4 mm×150 mm, 5 µm), detection wavelength:254 nm, mobile phase:water-methanol-perchloric acid (19:1:0.1), flow rate:0.75 ml/min, sample injection volume:10 µl. RESULTS: D-valaciclovir was completely separated from L-enantiomer impurity. The contents of L-enantiomer impurity were 0.65%-2.62% on average in 8 batches of valaciclovir hydrochloride. CONCLUSION: Enantiomeric impurity contents in each batch of products were all meet criteria of United States Pharmacopeia, which can be used in criteria of Chinese Pharmacopeia as references.


Assuntos
Aciclovir/análogos & derivados , Cromatografia Líquida de Alta Pressão/métodos , Valina/análogos & derivados , Aciclovir/análise , Estereoisomerismo , Valaciclovir , Valina/análise
12.
Mol Pharm ; 10(8): 3090-102, 2013 Aug 05.
Artigo em Inglês | MEDLINE | ID: mdl-23808658

RESUMO

The success of gene therapy largely relies on a safe and effective gene delivery system. The objective of this study is to design a highly efficient system for the transfection of epidermal stem cells (ESCs) and investigate the transfected ESCs (TESCs) as a therapeutic agent and gene delivery reservoir for wound treatment. As a nonviral vector, ß-cyclodextrin-linked polyethylenimines (CYD-PEI) was synthesized by linking ß-cyclodextrin with polyethylenimines (600 Da). Gelatin scaffold incorporating ß-tricalcium phosphate (ß-TCP) was utilized as a substrate for the culture and transfection of ESCs. With the CYD-PEI/pDNA-VEGF165 polyplexes incorporated gelatin/ß-TCP scaffold based 3D transfection system, prolonged VEGF expression with a higher level was obtained at day 7 in ESCs than those in two-dimensional plates. Topical application of the TESCs significantly accelerated the skin re-epithelization, dermal collagen synthesis, and hair follicle regeneration. It also exhibited a potential in scar inhibition by regulating the distribution of different types of collagen. In contrast to ESCs, an additive capacity in stimulating angiogenesis at the wound site was observed in the TESCs. The present study provides a basis for the TESCs as a promising therapeutic agent and gene delivery reservoir for wound therapy.


Assuntos
Fosfatos de Cálcio/química , Células Epidérmicas , Gelatina/química , Polietilenoimina/química , Células-Tronco/metabolismo , Fator A de Crescimento do Endotélio Vascular/genética , Cicatrização/efeitos da radiação , beta-Ciclodextrinas/química , Animais , Células Cultivadas , Técnicas de Transferência de Genes , Terapia Genética , Nanopartículas/química , Ratos , Ratos Sprague-Dawley , Células-Tronco/citologia , Células-Tronco/fisiologia , Cicatrização/fisiologia
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