RESUMEN
Tumor-associated macrophage (TAM) is regarded as an appealing cell target for cancer immunotherapy. However, it remains challenging to selectively eliminate M2-like TAM in tumor microenvironment. In this work, we employed a legumain-sensitive dual-coating nanosystem (s-Tpep-NPs) to deliver CSF-1R inhibitor pexidartinib (PLX3397) for targeting TAM therapy. The PLX3397-loaded NPs exhibited uniform size of â¼240 nm in diameter, good drug loading capacity and efficiency, as well as sustained drug release profile. Compared to non-sensitive counterpart ns-Tpep-NPs, s-Tpep-NPs showed distinguished selectivity upon M1 and M2 macrophage uptake with relation to incubation time and dose. Besides, the selectivity of anti-proliferation effect was also identified for s-Tpep-NPs against M1 and M2 macrophage. In vivo imaging demonstrated that s-Tpep-NPs exhibited much higher tumoral accumulation and TAM recognition specificity as compared to non-sensitive ns-Tpep-NPs. In vivo efficacy verified that s-Tpep-NPs formulation was much more effective than ns-Tpep-NPs and other PLX3397 formulations to treat B16F10 melanoma via targeting TAM depletion and modulating tumor immune microenvironment. Overall, this study provides a robust and promising nanomedicine strategy for TAM-targeted cancer immunotherapy.
Asunto(s)
Nanopartículas , Neoplasias , Macrófagos Asociados a Tumores , Línea Celular Tumoral , Inmunoterapia/métodos , Microambiente TumoralRESUMEN
Tumour-associated macrophages (TAMs) represent an attractive cell target for anticancer therapy. However, selective and efficient targeting of TAMs remains difficult. Here, we constructed a novel dually functionalised nanoparticle platform (s-Tpep-NPs) by surface co-modification of nanoparticles (NPs) with tuftsin (Tpep) and legumain protease-sheddable polyethylene glycol 5k (PEG5k) to achieve selective targeted delivery to TAMs. The fluorescence resonance energy transfer experiment and in vitro cellular uptake assay confirmed that s-Tpep-NPs can responsively shed PEG5k and transform into active Tpep-NPs upon the cleavage of legumain that is overexpressed on TAM surfaces, which then promotes TAM phagocytosis through Fc receptor-mediated pathways. Owing to the shielding effect by legumain-sheddable PEG5k, s-Tpep-NPs can effectively decrease the Tpep-induced non-specific accumulation in mononuclear phagocyte system (MPS) organs during systemic circulation. Moreover, s-Tpep-NPs can significantly enhance the tumoural accumulation and improve the specificity and efficiency of targeting to TAMs, as compared with both controls of Tpep-NPs and non-sheddable ns-Tpep-NPs. Overall, this study provides a robust nanoplatform with a novel avenue for improved selectivity of targeted delivery to TAMs.
Asunto(s)
Nanopartículas , Tuftsina , Cisteína Endopeptidasas , Péptido Hidrolasas , Polietilenglicoles , Macrófagos Asociados a TumoresRESUMEN
M2 tumor-associated macrophages (TAMs) play a pivotal role in cancer progression and therapy resistance. Inhibition of TAMs is of great significance to reshape the protumor environment to benefit therapeutic outcomes. In this work, we developed a novel TAMs and tumor cells dual-targeting nanoparticle (ATpep-NPs) system for cancer chemotherapy by integrating a docetaxel (DTX)-loaded nanocarrier and a multi-function peptide ATpep, which is composed of a phagocytosis-stimulating peptide-tuftsin (Tpep) fused with a substrate peptide-alanine-alanine-asparagine (AAN) of endoprotease legumain. In vitro protelytic and cellular uptake assays confirmed ATpep-NPs can be responsively activated into Tpep-NPs by cleavage of legumain that is overexpressed in both tumor cells and TAMs, which then promoted tumor cells internalization and TAMs phagocytosis through neuropilin-1/Fc receptor pathways. Due to AAN deactivation effect, ATpep-NPs can effectively decrease the Tpep-induced non-specific uptake by M1-polarized and normal macrophage during systemic circulation. Our results of in vivo experiments demonstrated ATpep-NPs outperformed Tpep-NPs in tumor and TAMs dual-targeting delivery efficiency with markedly enhanced efficacy against both tumor growth inhibition and TAMs depletion. Overall, this study offers a novel approach for development of multitargeted delivery vehicle for improved cancer chemotherapy.
Asunto(s)
Antineoplásicos , Nanopartículas , Neoplasias , Tuftsina , Antineoplásicos/farmacología , Línea Celular Tumoral , Cisteína Endopeptidasas , Péptido HidrolasasRESUMEN
Solid lipid nanoparticles (SLNs) have been widely used as a vehicle for drug delivery. However, highly ordered lipid lattices and poor storage stability limit their practical application. Highly ordered crystal lattices may result from the low drug payload. In addition, the lipid matrix of SLNs may undergo a polymorphic transition from high energy and disordered modifications to low energy and ordered modifications during storage. This leads to drug expulsion and precipitation. Meanwhile, SLNs are susceptible to particle aggregation and size growth during storage. To improve the performance of SLNs, two comb-shaped amphiphilic macromolecular materials (CAMs), dodecyl inulin (Inu12) and octadecyl inulin (Inu18), were synthesized and utilized as emulsifiers to modify and stabilize SLNs (Inu12/Inu18-SLNs). The results indicated that Inu12 and Inu18 could more effectively reduce the lipid crystallinity and crystal lattice order of fresh SLNs versus Poloxamer 188 and Tween-80. Moreover, after six months of storage at 4 °C or 25 °C, both blank and Cyclosporine A (CsA)-loaded Inu12/Inu18-SLNs had a slower crystal transition than Tween/P188-SLNs. The particle size increases of Inu12/Inu18-SLNs were much smaller than those of Tween/P188-SLNs. The drug encapsulation efficiencies of CsA-loaded Inu12/Inu18-SLNs during storage decreased more slowly than Tween-SLNs. Therefore, Inu12 and Inu18 could more effectively inhibit lipid crystal transition and prevent particle aggregation during storage. This, in turn, leads to better storage physical stability of SLNs. Thus, the Inu12 and Inu18 CAMs were superior to Tween-80 and Poloxamer 188 (common straight-chain surfactants).
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Inulina/química , Lípidos/química , Nanopartículas/química , Tensoactivos/química , Tamaño de la Partícula , Propiedades de SuperficieRESUMEN
Cancer stem cells (CSCs) can resist conventional chemotherapy to lead to cancer recurrence. For complete eradication of cancers, an effective CSCs therapeutic strategy should be developed to combine with conventional chemotherapy. In this work, a novel vitamin E-based redox-sensitive salinomycin (SAL, an inhibitor for CSCs) prodrug nanoparticles (TS NPs) and hyaluronic acid (HA)-coated TS NPs (HTS NPs) were fabricated to deliver paclitaxel (PTX) for cancer-targeted and combined chemotherapy. Both TS and HTS prodrug NPs had mean diameter of about 200 nm with uniform size distribution, excellent drug loading capacity for PTX, and glutathione-triggered SAL and PTX release profiles. The HTS prodrug NPs had enhanced cellular uptake efficiency over TS NPs due to CD44 receptor-mediated endocytosis, hence exerting stronger potency of SAL upon CSCs-enriched mammospheres formation and G0/G1 cell phase arresting. Cytotoxicity and 3D tumor spheroids assays demonstrated that both TS and HTS prodrug NPs themself can synergize with loaded PTX to maximize the chemotherapeutic effect. Obviously, the latter demonstrated a more potent anticancer efficacy due to improved intracellular drug delivery efficiency. These results suggested that the designed TS prodrug NPs, especially the coated HTS NPs can serve as an effective anti-CSCs strategy for cancer targeted and combination treatments.
Asunto(s)
Protocolos de Quimioterapia Combinada Antineoplásica/farmacología , Neoplasias de la Mama/tratamiento farmacológico , Nanopartículas/química , Paclitaxel/uso terapéutico , Profármacos/farmacología , Piranos/uso terapéutico , Vitamina E/farmacología , Neoplasias de la Mama/patología , Puntos de Control del Ciclo Celular/efectos de los fármacos , Muerte Celular/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Liberación de Fármacos , Femenino , Humanos , Células MCF-7 , Nanopartículas/ultraestructura , Células Madre Neoplásicas/patología , Oxidación-Reducción , Paclitaxel/farmacología , Tamaño de la Partícula , Profármacos/síntesis química , Profármacos/química , Piranos/síntesis química , Piranos/química , Piranos/farmacología , Esferoides Celulares/efectos de los fármacos , Esferoides Celulares/patología , Difracción de Rayos XRESUMEN
Glioblastoma multiforme (GBM) is the most common and deadly brain cancer, characterized by its aggressive proliferation to adjacent tissue and high recurrence rate. We studied the efficacy and related mechanisms of the combination of cyclopamine (Cyp, a Sonic-hedgehog pathway (Shh) inhibitor) and temozolomide (TMZ, the clinically most used chemotherapeutic agent) in anti-GBM treatment. The micellarized Cyp (MCyp) showed better performance than Cyp solution in inhibiting GBM cells proliferation (3.77-fold against U87 MG cells and 3.28-fold against DBTRG-05MG cells) and clonogenity (1.35-fold against U87 MG cells and 2.17-fold against DBTRG-05MG cells), and preferred behavior of inhibiting cell invasion, colony formation through attenuated Gli1 expression. In addition, combination of MCyp and TMZ exhibited synergistic cytotoxicity, correlating with their ability in inducing apoptosis and eliminating neurospheres formation, and the combination of TMZ was accompanied with the enhanced blockage of Shh pathway. The optimal ratio of MCyp combined to TMZ was 1:20. So we proposed to use TMZ to kill tumor parenchyma and MCyp as the cancer stem cells inhibitor to resist tumor recurrence. These findings demonstrated that combination of TMZ with micellarized Cyp is a promising strategy for exerting different functions of drugs for tumor treatment.
Asunto(s)
Neoplasias Encefálicas/metabolismo , Dacarbazina/análogos & derivados , Glioblastoma/metabolismo , Micelas , Alcaloides de Veratrum/administración & dosificación , Proteína con Dedos de Zinc GLI1/metabolismo , Apoptosis/efectos de los fármacos , Neoplasias Encefálicas/tratamiento farmacológico , Neoplasias Encefálicas/genética , Neoplasias Encefálicas/patología , Línea Celular Tumoral , Movimiento Celular/efectos de los fármacos , Proliferación Celular/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Dacarbazina/administración & dosificación , Sinergismo Farmacológico , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Glioblastoma/tratamiento farmacológico , Glioblastoma/genética , Glioblastoma/patología , Humanos , Transducción de Señal/efectos de los fármacos , Temozolomida , Ensayo de Tumor de Célula Madre , Proteína con Dedos de Zinc GLI1/genéticaRESUMEN
Metastasis of cancer makes up the vast majority of cancer-related deaths, and it usually initiates from tumor cells invasiveness and develops through tumor neovasculature. In this work, we have fabricated a CD44/neuropilin dual receptor-targeting nanoparticulate system (tLyP-1-HT NPs) with endogenous or FDA approved components for treating metastatic triple negative breast cancer (TNBC). The enhanced specific targeting of tLyP-1-HT NPs to both metastatic tumor cells and metastasis-supporting tumor neovasculature was contributed by means of CD44/neuropilin dual receptor-mediated interaction. The NPs not only effectively suppress the invasive capability of tumor cells themselves, but also significantly restrain the metastasis incidence via extravasation as well as the eventual colonization in lungs. In all the three types of TNBC-bearing mice models, orthotopic, post-metastasis and metastasis prevention models, the docetaxel-loaded tLyP-1-HT NPs exhibited markedly enhanced anti-tumor and anti-metastasis efficacy. The inhibitory rates of tLyP-1-HT NPs against orthotopic tumor growth and lung metastasis achieved 79.6% and 100%, respectively. The metastasis inhibition rate and life extension rate of the tLyP-1-HT NPs against post-pulmonary metastasis mice reached 85.1% and up to 62.5%, respectively. All the results demonstrated the designed dual receptor-targeting multifunctional NPs hold great potential in treating metastatic TNBC and lung metastasis.
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Receptores de Hialuranos/metabolismo , Nanopartículas/química , Neuropilinas/metabolismo , Neoplasias de la Mama Triple Negativas/tratamiento farmacológico , Animales , Antineoplásicos/uso terapéutico , Línea Celular Tumoral , Docetaxel , Portadores de Fármacos , Liberación de Fármacos , Femenino , Humanos , Neoplasias Pulmonares/irrigación sanguínea , Neoplasias Pulmonares/tratamiento farmacológico , Neoplasias Pulmonares/secundario , Ratones Endogámicos BALB C , Ratones Desnudos , Terapia Molecular Dirigida , Tamaño de la Partícula , Polímeros/química , Propiedades de Superficie , Taxoides/uso terapéutico , Neoplasias de la Mama Triple Negativas/irrigación sanguínea , Neoplasias de la Mama Triple Negativas/metabolismo , Neoplasias de la Mama Triple Negativas/patologíaRESUMEN
PROBLEM: We aimed to investigate the modulatory effects of vitamin D on peripheral blood cellular immune response in patients with recurrent miscarriage (RM). METHOD OF STUDY: The effect of vitamin D on the number of peripheral blood cells, T helper 1 (Th1) cytokines, and NK cytotoxicity was measured in 99 women with RM. RESULTS: The percentage of CD19+ B cells and NK cytotoxicity at an effector-to-target cell (E:T) ratio of 50:1, 25:1, and 12.5:1 were significantly higher in the vitamin D insufficiency group (VDI) than in the vitamin D normal group (VDN) (P<.05 each). The proportion of TNF-α-expressing Th cells was significantly higher in the vitamin D deficiency group (VDD) than in VDN (P<.05). However, there were no significant differences between VDI and VDD. This dysregulation was significantly reduced with 1,25(OH)2 D supplementation. CONCLUSION: The data suggest that the abnormalities of cellular immune response were observed in RM patients with a low vitamin D level, which could be regulated to some extent with 1,25(OH)2 D supplementation.
Asunto(s)
Aborto Habitual/inmunología , Calcitriol/administración & dosificación , Factores Inmunológicos/administración & dosificación , Deficiencia de Vitamina D/inmunología , Aborto Habitual/dietoterapia , Aborto Habitual/genética , Aborto Habitual/patología , Administración Oral , Adulto , Linfocitos B/efectos de los fármacos , Linfocitos B/inmunología , Linfocitos B/patología , Estudios de Casos y Controles , Citotoxicidad Inmunológica , Femenino , Expresión Génica , Humanos , Inmunofenotipificación , Interferón gamma/genética , Interferón gamma/inmunología , Células Asesinas Naturales/efectos de los fármacos , Células Asesinas Naturales/inmunología , Células Asesinas Naturales/patología , Embarazo , Células TH1/efectos de los fármacos , Células TH1/inmunología , Células TH1/patología , Factor de Necrosis Tumoral alfa/genética , Factor de Necrosis Tumoral alfa/inmunología , Deficiencia de Vitamina D/dietoterapia , Deficiencia de Vitamina D/genética , Deficiencia de Vitamina D/patologíaRESUMEN
Low density lipoprotein (LDL), which is a principal carrier for the delivery of cholesterol, has been used as a great candidate for the delivery of drugs to tumor based on the great requirements for cholesterol of many cancer cells. Mimicking the structure and composition of LDL, we designed a synthetic low-density lipoprotein (sLDL) to encapsulate paclitaxel-alpha linolenic acid (PALA) for tumor therapy. The PALA loaded sLDL (PALA-sLDL) and PALA-loaded microemulsion (PALA-ME, without the binding domain for LDLR) displayed uniform sizes with high drug loading efficiency (> 90%). In vitro studies demonstrated PALA-sLDL exhibited enhanced cellular uptake capacity and better cytotoxicity to LDLR over-expressed U87 MG cells as compared to PALA-ME. The uptake mechanisms of PALA-sLDL were involved in a receptor mediated endocytosis and macropinocytosis. Furthermore, the in vivo biodistribution and tumor growth inhibition studies of PALA-sLDL were investigated in xenograft U87 MG tumor-bearing mice. The results showed that PALA-sLDL exhibited higher tumor accumulation than PALA-ME and superior tumor inhibition efficiency (72.1%) compared to Taxol® (51.2%) and PALA-ME (58.8%) but with lower toxicity. These studies suggested that sLDL is potential to be used as a valuable carrier for the selective delivery of anticancer drugs to tumor with low systemic toxicity.
Asunto(s)
Portadores de Fármacos , Lipoproteínas LDL , Neoplasias/tratamiento farmacológico , Paclitaxel/administración & dosificación , Paclitaxel/efectos adversos , Animales , Antineoplásicos/administración & dosificación , Antineoplásicos/efectos adversos , Línea Celular Tumoral , Portadores de Fármacos/efectos adversos , Portadores de Fármacos/farmacocinética , Sistemas de Liberación de Medicamentos/efectos adversos , Células Hep G2 , Humanos , Lipoproteínas LDL/administración & dosificación , Lipoproteínas LDL/farmacocinética , Masculino , Ratones , Ratones Endogámicos BALB C , Ratones Desnudos , Micelas , Neoplasias/metabolismo , Tamaño de la Partícula , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Marinesco-Sjögren syndrome (MSS) is a rare autosomal recessive disorder, which is characterized by congenital cataracts, cerebellar ataxia, progressive muscle weakness, and delayed psychomotor development. SIL1, which is located at 5q31.2, is the only gene known to cause MSS. Dandy-Walker syndrome (DWS) is defined by hypoplasia, upward rotation of the cerebellar vermis, and cystic dilation of the fourth ventricle; however, its genetic pathogeny remains unclear. Here, we report a Chinese consanguineous family with MSS and DWS. Whole exome sequencing identified a novel nonstop mutation in SIL1. Sanger sequencing revealed that the mutation was segregated in this family according to a recessive mode of inheritance. We found that the mutation changed a stop codon (TGA) to an arginine codon (CGA), and no in-frame termination codon in the 3' untranslated region (UTR) of SIL1 could be found. The mRNA levels of SIL1 were decreased by 56.6% and 37.5% in immortalized lymphoblasts of the patients respectively; the protein levels of SIL1 were substantially decreased. This case study is the first report on Chinese MSS patients, MSS complicated by DWS, and a nonstop mutation in SIL1. Our findings imply the pathogenetic association between DWS and MSS.
Asunto(s)
Pueblo Asiatico/genética , Consanguinidad , Síndrome de Dandy-Walker/complicaciones , Síndrome de Dandy-Walker/genética , Factores de Intercambio de Guanina Nucleótido/genética , Mutación , Degeneraciones Espinocerebelosas/complicaciones , Degeneraciones Espinocerebelosas/genética , Adolescente , Niño , China , Exoma/genética , Femenino , Factores de Intercambio de Guanina Nucleótido/análisis , Factores de Intercambio de Guanina Nucleótido/biosíntesis , Humanos , Masculino , LinajeRESUMEN
Poor site-specific delivery and incapable deep-penetration into tumor are the intrinsic limitations to successful chemotherapy. Here, the tumor-homing penetrating peptide tLyP-1-functionalized nanoparticles (tLPTS/HATS NPs), composed of two modularized amphiphilic conjugates of tLyP-1-PEG-TOS (tLPTS) and TOS-grafted hyaluronic acid (HATS), had been fabricated for tumor-targeted delivery of docetaxel (DTX). The prepared tLPTS/HATS NPs had about 110 nm in mean diameter, high drug encapsulation efficiency (93%), and sustained drug release behavior. In vitro studies demonstrated that the tLPTS/HATS NPs exhibited enhanced intracellular delivery and much better anti-invasion ability, cytotoxicity, and apoptosis against both invasive PC-3 and MDA-MB-231 cells as compared to the non-tLyP-1-functionalized HATS NPs. The remarkable penetrability and inhibitory effect on both PC-3 and MDA-MB-231 multicellular tumor spheroids were also identified for the tLPTS/HATS NPs. In vivo biodistribution imaging demonstrated the tLPTS/HATS NPs possessed much more lasting accumulation and extensive distribution throughout tumor regions than the HATS NPs. The higher in vivo therapeutic efficacy with lower systemic toxicity of the tLPTS/HATS NPs was also verified by the PC-3 xenograft model in athymic nude mice. These results suggested that the designed novel tLPTS/HATS NPs were endowed with tumor recognition, internalization, penetration, and anti-invasion, and thus might be a promising anticancer drug delivery vehicle for targeted cancer therapy.
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Antineoplásicos/administración & dosificación , Ácido Hialurónico/química , Nanopartículas , Neoplasias/tratamiento farmacológico , Péptidos/metabolismo , alfa-Tocoferol/química , Animales , Línea Celular Tumoral , Humanos , Masculino , Ratones , Ratones Endogámicos BALB C , Neoplasias/metabolismo , Neoplasias/patología , Péptidos/químicaRESUMEN
There are several obstacles in the process of successful treatment of malignant tumors, including toxicity to normal cells, inefficiency of drug permeation and accumulation into the deep tissue of solid tumor, and multidrug resistance (MDR). In this work, we prepared docetaxel (DTX)-loaded hybrid micelles with DSPE-PEG and TPGS (TPGS/DTX-M), where TPGS serves as an effective P-gp inhibitor for overcoming MDR, and active targeting hybrid micelles (FA@TPGS/DTX-M) with targeting ligand of folate on the hybrid micelles surface offering active targeting to folate receptor-overexpressed tumor cells. A systematic comparative evaluation of these micelles on cellular internalization, sub-cellular distribution, antiproliferation, mitochondrial membrane potential, cell apoptosis and cell cycle, permeation and inhibition on 3-dimensional multicellular tumor spheroids, as well as antitumor efficacy and safety assay in vivo were well performed between sensitive KB tumors and resistant KBv tumors, and among P-gp substrate or not. We found that the roles of folate and TPGS varied due to the sensitivity of tumors and the loaded molecules in the micelles. Folate and folate receptor-mediated endocytosis played a leading role in internalization, permeation and accumulation for sensitive tumors and non-substrates of P-gp. On the contrary, TPGS played the predominant role which dramatically decreased the efflux of drugs both when the tumor is resistant and for P-gp substrate. These findings are very meaningful for guiding the design of carrier delivery system to treat tumors. The antitumor efficacy in xenograft nude mice model and safety assay showed that the TPGS/DTX-M and FA@TPGS/DTX-M significantly exhibited higher antitumor activity against resistant KBv tumors than the marketed formulation and normal micelles owing to the small size (approximately 20 nm), hydrophilic PEGylation, TPGS inhibition of P-gp function, and folate receptor-modified endocytosis, permeation and accumulation in solid tumor, as well as synergistic effects of DTX-induced cell division inhibition, growth restraint and TPGS-triggered mitochondrial apoptosis in tumor cells. In conclusion, folate-modified TPGS hybrid micelles provide a synergistic strategy for effective delivery of DTX into KBv cells and overcoming MDR.
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Resistencia a Múltiples Medicamentos , Resistencia a Antineoplásicos , Micelas , Vitamina E/análogos & derivados , Miembro 1 de la Subfamilia B de Casetes de Unión a ATP/antagonistas & inhibidores , Animales , Antineoplásicos/administración & dosificación , Antineoplásicos/uso terapéutico , Proliferación Celular/efectos de los fármacos , Docetaxel , Femenino , Ligandos , Potencial de la Membrana Mitocondrial/efectos de los fármacos , Ratones , Ratones Endogámicos BALB C , Polietilenglicoles/farmacología , Taxoides/administración & dosificación , Taxoides/uso terapéutico , Vitamina E/farmacologíaRESUMEN
OBJECTIVE: To detect the underlying genetic defect in two Chinese families with hereditary multiple exostoses and provide genetic counseling. METHODS: Potential mutations in EXT1 and EXT2 genes in the probands were detected by direct sequencing of PCR-amplified exons. Suspected mutations were verified in all available family members and 200 unrelated healthy controls. RESULTS: A heterozygous frameshift mutation c.346_356delinsTAT in exon 1 of EXT1 and a heterozygous deletion mutation c.2009-2012del(TCAA) in exon 10 of EXT1 were respectively detected in affected members from the two families. The same mutations were not detected in unaffected members and 200 unrelated healthy controls. No mutations in EXT2 were detected in the two families. CONCLUSION: Two novel mutations of EXT1 have been detected in association with hereditary multiple exostoses in two Chinese families. Above results have provided a basis for genetic counseling for the two families and expanded the spectrum of EXT1 mutations.
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Exostosis Múltiple Hereditaria/enzimología , Exostosis Múltiple Hereditaria/genética , N-Acetilglucosaminiltransferasas/genética , Eliminación de Secuencia , Adolescente , Adulto , Anciano , Pueblo Asiatico/genética , Niño , Preescolar , Análisis Mutacional de ADN/métodos , Femenino , Heterocigoto , Humanos , Masculino , Persona de Mediana Edad , Linaje , Adulto JovenRESUMEN
Currently, there are difficulties associated with the culturing of pluripotent human embryonic stem cells (hESCs), and knowledge regarding their regulatory mechanisms is limited. MicroRNAs (miRNAs) regulate gene expression and have critical functions in stem cell self-renewal and differentiation. Moreover, fibroblast growth factor (FGF) and the insulin-like growth factor receptor (IGF-1R) are key activators of signaling in hESCs. Based on the identification of complementary binding sites in miR-223 and IGF-1R mRNA, it is proposed that miR-223 acts as a local regulator of IGF-1R. Therefore, levels of miR-223 were detected in differentiated versus undifferentiated hESCs. In addition, proliferation, apoptosis, and differentiation were assayed in these two hESC populations and were compared in the presence of exogenous miR-223 and miR-223 inhibitor. Inhibition of miR-223 was found to maintain the undifferentiated state of hESCs, while addition of miR-223 induced differentiation. Furthermore, these effects were found to be likely dependent on IGF-1R/Akt signaling.
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Diferenciación Celular/genética , MicroARNs/genética , Proteínas Proto-Oncogénicas c-akt/genética , Receptor IGF Tipo 1/genética , Apoptosis/genética , Línea Celular , Proliferación Celular/genética , Células Madre Embrionarias , Factores de Crecimiento de Fibroblastos/genética , Regulación del Desarrollo de la Expresión Génica , Humanos , MicroARNs/antagonistas & inhibidores , Proteínas Proto-Oncogénicas c-akt/biosíntesis , ARN Mensajero/genética , Receptor IGF Tipo 1/biosíntesis , Transducción de Señal/genéticaRESUMEN
OBJECTIVE: To perform genetic analysis in 5 patients with blepharophimosis-ptosis-epicanthus inversus syndrome (BPES) and refine the genotype-phenotype correlation. METHODS: G-band karyotyping, fluorescent in situ hybridization (FISH), SNP array, PCR and sequencing techniques were performed to one patient with BPES and mental retardation and 4 only with BPES. RESULTS: Patient 1 with mental retardation carried a 9.4 Mb heterozygous deletion in chromosome 3q22.1-q23 including FOXL2 gene; Both patient 2 and 3 carried a c.704delG heterozygous mutation of FOXL2, while they were assigned to the different clinical type from those reported previously. Patient 3 was assigned to type II BPES; No mutation of FOXL2 was detected in patient 4 and 5. CONCLUSIONS: There might be the gene(s) responsible for mental retardation within chromosome 3q22.1-q23. It was indicated that the mutation c.704delG in FOXL2 led to a truncated protein is associated with both type I and II of BPES.
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Blefarofimosis/genética , Factores de Transcripción Forkhead/genética , Mutación , Adolescente , Adulto , Niño , Preescolar , Femenino , Proteína Forkhead Box L2 , Humanos , Lactante , Masculino , Linaje , Polimorfismo de Nucleótido Simple , Eliminación de Secuencia , Síndrome , Adulto JovenRESUMEN
OBJECTIVE: To evaluate the conventional cytogenetic methods in genetic diagnosis and prenatal diagnosis in the family with a proband of Angelman syndrome (AS). METHODS: High-resolution G-banding karyotyping and fluorescence in situ hybridization (FISH) on metaphase chromosomes were performed. RESULTS: Two AS patients and 1 normal fetus in the family were successfully detected by FISH. CONCLUSION: Our result demonstrated that patient with type I AS could be detected by combining the techniques of high-resolution G-banding and FISH with clinical observation, which would offer accurate genetic counseling information to the geneticists and provide the prenatal diagnosis for the AS family.
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Síndrome de Angelman/diagnóstico , Síndrome de Angelman/genética , Diagnóstico Prenatal , Adulto , Preescolar , Cromosomas Humanos Par 15/genética , Femenino , Humanos , Hibridación Fluorescente in Situ , Lactante , Cariotipificación , Masculino , EmbarazoRESUMEN
OBJECTIVE: To develop a rapid genetic diagnosis technique for the patients with hereditary hearing loss by screening hot spots of mutations, namely 235delC of the GJB2 gene, IVS7-2A>G of the SLC26A4 gene, and 1555A>G of mitochondrial 12S rRNA. METHODS: Multiple PCR amplification of the three fragments covering the expected mutations in GJB2, SLC26A4 and 12S were carried out and the amplified products were analyzed by restriction fragment length polymorphism (RFLP). RESULTS: Eighteen homozygous and 18 heterozygous 235delC, 2 homozygous and 13 heterozygous IVS7-2A>G, and 8 homogeneous 1555A>G were detected in the 200 patients with hearing loss. All the results were confirmed by sequencing. The detection rate of the three mutant alleles was 21.7% (71/400 + 8/200 = 0.217) and the genetic diagnosis rate was 14% [(18+2+8)/200 = 0.14]. CONCLUSION: It is a convenient, efficient and economical method to screen the hot spots of mutation in the patient with hereditary hearing loss by using PCR-RFLP.
Asunto(s)
Pueblo Asiatico/genética , Pérdida Auditiva/genética , Mutación , Reacción en Cadena de la Polimerasa/métodos , Polimorfismo de Longitud del Fragmento de Restricción , Adolescente , Adulto , Niño , Preescolar , Conexina 26 , Conexinas/genética , Femenino , Humanos , Masculino , Proteínas de Transporte de Membrana/genética , Transportadores de Sulfato , Adulto JovenRESUMEN
OBJECTIVE: Mucopolysaccharidosis type II (MPSII) is a lethal, X-linked recessive disorder caused by mutation of iduronate-2-sulfatase (IDS) gene. Up to now there is no really effective treatment for this disorder, therefore it is important to provide an accurate genetic diagnosis and prenatal diagnosis for the MPSII families. In this study, we identify the pathogenic mutation in a Chinese family with MPSII. METHOD: The 8 years old male proband from a Chinese family was clinically diagnosed with MPSII. There are other 4 patients with similar phenotypes in the family who died at 9, 11, 7 and 10 years of age, respectively. Mutation analysis was carried out by polymerase chain reaction and direct sequencing of all exons and exon/intron boundaries of IDS gene. Denaturing high performance liquid chromatography (DHPLC) analysis was performed to screen the unknown variations of IDS gene in 100 unrelated control males. RESULT: Two allelic variants of exon 5 (c.684A > G) and exon 6 (c.851C > T) and a nonsense mutation of exon 7 (c.892C > T) were detected in IDS gene of the proband. Heterozygous mutations c.684A > G, c.851C > T and c.892C > T were detected in both proband's mother and maternal grandmother. The unknown variations of c.684A > G and c.851C > T were not found in the 100 unrelated control males. The male fetus (IV11) inherited the same mutation of IDS gene as the proband. CONCLUSION: Mutation c.892C > T of IDS gene causes MPSII in this family and prenatal diagnosis in one affected fetus was achieved.
Asunto(s)
Iduronato Sulfatasa/genética , Mucopolisacaridosis II/diagnóstico , Mucopolisacaridosis II/genética , Adolescente , Adulto , Pueblo Asiatico/genética , Niño , Análisis Mutacional de ADN , Familia , Femenino , Humanos , Masculino , Persona de Mediana Edad , Mutación , Fenotipo , Embarazo , Diagnóstico PrenatalRESUMEN
OBJECTIVE: To study the SLC26A4 gene mutations in patients with nonsyndromic hearing loss (NSHL) and provide the clinical guidance of gene diagnosis. METHODS: PCR and denaturing high-performance liquid chromatography (DHPLC) were used to screen the 21 exons and their flanking regions of the SLC26A4 gene. Samples with abnormal DHPLC wave patterns were sequenced to identify the variations. RESULTS: Among the 30 unrelated NSHL patients in whom no deafness-causing mutations of the GJB2 gene were identified, 10 types of variations were detected, including 7 known mutations, 2 novel mutations (F572L and D87Y), and 1 known polymorphism (Ivs11+47T>C). The Ivs7-2A>G is the most common type of variation, accounting for 40% of all the mutations. CONCLUSION: SLC26A4 mutation is a major cause of NSHL, just next to the GJB2 mutations. For NSHL patients without deafness-causing GJB2 mutations, the SLC26A4 mutation rate was 23.3%, and the Ivs7-2A>G was the most common mutation.