RESUMO
X-linked hypohidrotic ectodermal dysplasia (XLHED; OMIM 305100) is the most common form of ectodermal dysplasia, presenting with the triad of hypotrichosis, hypodontia, and hypohidrosis. This disorder is caused by mutations in EDA, which encodes ectodysplasin A, a member of the tumor necrosis factor superfamily. In this study, we describe clinical and genetic characteristics of 10 Korean XLHED patients (9 males, 1 female) from 9 families. Nine out of the 10 patients manifested the cardinal triad of symptoms. Six patients had a positive family history, while 2 patients were brothers. The most common initial presentation was hypotrichosis or hypodontia, while 1 patient presented with recurrent high fever in early infancy. Sanger sequencing of the EDA gene was performed and revealed 9 different mutations. Three had been reported previously, and 6 were novel mutations. One female patient, carrying a previously reported missense mutation, might be affected by skewed X-inactivation. This is the first observational study investigating genetically confirmed XLHED patients in Korea. To provide appropriate supportive care and genetic counseling, clinicians should consider the possibility of XLHED in the differential diagnosis of recurrent fever in infants, as well as recognize the typical triad of symptoms.
Assuntos
Displasia Ectodérmica Anidrótica Tipo 1/genética , Ectodisplasinas/genética , Mutação , Adolescente , Substituição de Aminoácidos , Criança , Pré-Escolar , Displasia Ectodérmica Anidrótica Tipo 1/etnologia , Ectodisplasinas/química , Feminino , Mutação da Fase de Leitura , Estudos de Associação Genética , Triagem de Portadores Genéticos , Humanos , Lactente , Masculino , Mutação de Sentido Incorreto , Linhagem , República da Coreia/epidemiologiaRESUMO
This study aimed to verify, in in vivo settings, whether quorum-sensing inhibition molecules could attenuate alveolar bone loss induced by Porphyromonas gingivalis/Fusobacterium nucleatum co-infection and reduce the bacterial colonization of periodontal tissues. In BALB/c mice, periodontitis was induced through oral inoculation with P. gingivalis and F. nucleatum six times during a 42-d period. Quorum sensing inhibitors (a furanone compound and D-ribose) were administered simultaneously with bacterial infection. Linear and volumetric modifications of interproximal alveolar bone levels were compared between groups using micro-computed tomography. Total bacteria, and P. gingivalis and F. nucleatum DNA in periodontal tissues, were quantified using real-time PCR. Radiographic linear measurements demonstrated a significant reduction of alveolar bone loss, of approximately 40%, in mice treated with quorum sensing inhibitors when compared with the co-infection group. This was confirmed by a significant increase of residual bone volume in the test group. While total bacterial genes in the treatment group significantly decreased by 93% in periodontal tissue samples when quorum sensing inhibitors were administered, no significant differences of P. gingivalis DNA were found. Quorum sensing inhibitors reduced periodontal breakdown and bacterial infection in periodontal tissues after co-infection with P. gingivalis and F. nucleatum.
Assuntos
Coinfecção , Periodontite , Percepção de Quorum/efeitos dos fármacos , Perda do Osso Alveolar , Animais , DNA Bacteriano/análise , Modelos Animais de Doenças , Furanos/administração & dosagem , Furanos/antagonistas & inibidores , Fusobacterium nucleatum/genética , Fusobacterium nucleatum/patogenicidade , Expressão Gênica , Genes Bacterianos , Interações Hospedeiro-Patógeno , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Periodontite/diagnóstico por imagem , Periodontite/microbiologia , Periodontite/patologia , Porphyromonas gingivalis/genética , Porphyromonas gingivalis/patogenicidade , Ribose/administração & dosagem , Ribose/antagonistas & inibidores , Microtomografia por Raio-XRESUMO
OBJECTIVE: To describe the characteristic MRI appearance of squatting-induced rhabdomyolysis involving the thigh muscles. METHODS: This study consisted of 10 cases obtained at 3 institutions from 2005 to 2015. A retrospective review was performed to obtain clinical information and MR scans for rhabdomyolysis of the thigh muscles. MRI was analyzed according to the distribution and degree of muscle involvement; the degree was assessed and graded as normal, mild or prominent. RESULTS: The mean patient age was 20.2 years (range, 15-24 years), and 7 of the 10 patients were male. All patients had history of excessive squatting action, suffered clinically from bilateral thigh pain and were confirmed to have rhabdomyolysis through analysis of serum creatine kinase (CK) levels. All of the patients (10/10) exhibited diffuse mild to prominent degree involvement of the anterior thigh muscles according to fluid-sensitive MR sequences. Among the anterior thigh muscles, the rectus femoris was spared in 8 patients (8/10) and mild degree involved in 2 patients (2/10). Thus, no cases exhibited prominent degree involvement of the rectus femoris muscle. CONCLUSION: Preservation of the rectus femoris muscle on MRI in squatting-induced rhabdomyolysis may be useful for differentiating rhabdomyolysis from other aetiologies. Advances in knowledge: Preservation of rectus femoris on MRI is distinguishable finding in squatting-induced rhabdomyolysis and reflects the functional anatomy of anterior thigh muscles.
Assuntos
Exercício Físico , Imageamento por Ressonância Magnética/métodos , Rabdomiólise/diagnóstico por imagem , Coxa da Perna/diagnóstico por imagem , Adolescente , Adulto , Feminino , Humanos , Masculino , Estudos Retrospectivos , Rabdomiólise/patologia , Coxa da Perna/patologia , Adulto JovemRESUMO
In this study, hMSCs encapsulated in a fibrin hydrogel containing heparinized NPs loaded with TGF-ß3 (100 ng/ml), or TGF-ß3 (100 ng/ml) alone, were subjected to growth factor release and denaturation tests at one, two and four weeks in in vitro culture systems. Additionally, stem cell differentiation was assessed via RT-PCR, real-time quantitative PCR (qPCR), histology, and immunohistochemical assays. In the in vivo studies with nude mouse, when transplanted into nude mice, hMSCs embedded in fibrin hydrogels survived and proliferated more readily in those samples containing TGF-ß3-loaded NPs, or TGF-ß3 alone, compared to those containing only NPs or the fibrin hydrogel alone. Additionally, RT-PCR, real-time qPCR, histology, Western blotting, and immunohistochemistry analyses revealed that chondrocyte-specific extracellular matrix (ECM) genes and their proteins were expressed at high levels by hMSCs embedded in hydrogels containing TGF-ß3-loaded NPs. Finally, the results observed in the rabbit animal model treated with hMSCs embedded in a fibrin hydrogel containing TGF-ß3-loaded NPs were also evaluated by the RT-PCR, real-time qPCR, histology, Western blotting, and immunohistochemistry analyses. The in vitro and in vivo results indicated that transplanted hMSCs together with TGF-ß3 may constitute a clinically efficient method for the regeneration of hyaline articular cartilage.
Assuntos
Condrogênese/fisiologia , Fibrina/química , Células-Tronco Mesenquimais/citologia , Animais , Western Blotting , Cartilagem Articular/cirurgia , Células Cultivadas , Feminino , Citometria de Fluxo , Humanos , Hidrogel de Polietilenoglicol-Dimetacrilato/química , Imuno-Histoquímica , Células-Tronco Mesenquimais/metabolismo , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Nanopartículas/química , Coelhos , Engenharia Tecidual/métodosRESUMO
The induction of stem cell differentiation by drugs and growth factors has been the objective of many studies designed to develop methods for the formation of new tissues or the repair of degenerated tissues via transplantation. In this study, drugs and growth factors with high potential for use in tissue repair were embedded in human mesenchymal stem cells (hMSCs), which were then induced to differentiate into chondrogenic, osteogenic, and adipogenic lineages. Additionally, microspheres coated and loaded with the drugs and growth factors successfully proliferated and, as expected, induced the differentiation of transplanted hMSCs into the desired specific cell types. Furthermore, hMSCs transplanted in micro-typed scaffolds prevented changes in differentiation. RT-PCR and western blot analysis of the resultant cartilage, bone, and adipose tissues showed that a combination of drugs and different growth factor types induced the differentiation of transplanted hMSCs. Additionally, histology and immunohistochemistry showed that specific ECMs and proteins released from transplanted hMSCs were present within the nanosphere-coated microspheres. The results of this study show that the regulation of stem cell differentiation by drugs and growth factors might enable the fabrication of therapeutic materials for the delivery of stem cells that are simpler to use, less expensive, and more easily controlled than the delivery systems currently available.
Assuntos
Adipogenia/efeitos dos fármacos , Condrogênese/efeitos dos fármacos , Peptídeos e Proteínas de Sinalização Intercelular/farmacologia , Células-Tronco Mesenquimais/citologia , Microesferas , Nanosferas/química , Osteogênese/efeitos dos fármacos , Fosfatase Alcalina/metabolismo , Animais , Adesão Celular/efeitos dos fármacos , Células Cultivadas , Materiais Revestidos Biocompatíveis/farmacologia , DNA/metabolismo , Imunofluorescência , Perfilação da Expressão Gênica , Regulação da Expressão Gênica/efeitos dos fármacos , Glicosaminoglicanos/metabolismo , Humanos , Ácido Láctico/farmacologia , Células-Tronco Mesenquimais/efeitos dos fármacos , Células-Tronco Mesenquimais/enzimologia , Células-Tronco Mesenquimais/ultraestrutura , Camundongos , Camundongos Nus , Nanosferas/ultraestrutura , Ácido Poliglicólico/farmacologia , Copolímero de Ácido Poliláctico e Ácido Poliglicólico , Alicerces Teciduais/químicaRESUMO
Micro-structured scaffolds formed with poly(lactic- co -glycolic acid) (PLGA) microspheres were composed of adhesion molecules and growth factors. PLGA microspheres, constructed with Arg-Gly-Asp (RGD) peptide and bone morphogenic protein 2 (BMP-2) were created as a stem cell delivery vehicle. In this study, a high potential for cell adhesion and differentiation of human mesenchymal stem cells (hMSCs) was achieved by constructing the scaffolds with different compositions of coating materials. Specific gene and protein detection by RT-PCR and western blot analysis of the embedded hMSCs revealed that a combination of RGD peptide and BMP-2 induced differentiation of bone cells. Histology and immunohistochemistry results confirmed that bone cell-differentiated transplanted hMSCs were present in the micro-structured scaffolds. The results of this study demonstrate that the regulation of stem cell differentiation by adhesion molecules and growth factors has the potential to enable formation of therapeutic vehicles for the delivery of stem cells that are easily fabricated, less expensive, and more easily controlled than currently available delivery systems. The micro-structure typed PLGA microspheres used in this study possessed unique properties of ideal scaffolds. The embedded hMSCs easily adhered onto the PLGA microspheres mediated by RGD peptide, proliferated well onto the scaffolds, and differentiated to perform the distinct functions of bone tissues.
Assuntos
Proteína Morfogenética Óssea 2/farmacologia , Diferenciação Celular/efeitos dos fármacos , Células-Tronco Mesenquimais/citologia , Células-Tronco Mesenquimais/efeitos dos fármacos , Microesferas , Oligopeptídeos/farmacologia , Osteogênese/efeitos dos fármacos , Animais , Células Cultivadas , Materiais Revestidos Biocompatíveis/farmacologia , Dexametasona/farmacologia , Humanos , Ácido Láctico/farmacologia , Camundongos , Camundongos Nus , Microscopia Confocal , Ácido Poliglicólico/farmacologia , Copolímero de Ácido Poliláctico e Ácido Poliglicólico , Propriedades de Superfície/efeitos dos fármacosRESUMO
Human mesenchymal stem cells (hMSCs) represent a potent target for gene delivery for both stem cell differentiation applications and clinical therapies. However, it has, thus far, proven difficult to develop delivery vehicles that increase the efficiency of gene delivery to hMSCs, due to several problematic issues. We have evaluated different vehicles with regard to the efficiency with which they deliver hMSCs and enhance the ability to deliver a reporter gene. In this study, a non-viral gene delivery system using nanoparticles was designed, with emphasis placed on the ability of the system to mediate high levels of gene expression into stem cells. Via polyplexing with polyethylenimine (PEI), the cell-uptake ability of the nanoparticles was enhanced for both in vitro and in vivo culture systems. In experiments with PEI/pNDA polyplexed with nanoparticles, the expression of green fluorescent protein (GFP) with this vehicle was noted in up to 75% of hMSCs 2 days after transfection, and GFP gene expression was detected via Western blotting, flow cytometric analysis, and immunofluorescence using a confocal laser microscope after transfection.
Assuntos
DNA/administração & dosagem , Vetores Genéticos , Células-Tronco Mesenquimais/metabolismo , Nanopartículas , Adulto , Animais , Western Blotting , Terapia Genética , Humanos , Masculino , Camundongos , Microscopia Eletrônica de TransmissãoRESUMO
This study was conducted to reveal the chondrogenesis of mesenchymal stem cells that had been genetically modified with the green fluorescence protein (GFP) gene and then co-cultured with chondrocytes in vitro and in vivo. Subsequent mixing of chondrocytes in the hydrogel constructs induced increased chondrogenic differentiation of the transfected hMSCs. The proliferation and differentiation of MSCs that were transfected with the GFP gene and co-cultured with chondrocytes (1:1 and 1:3) or chondrocytes alone were evaluated by a live/dead assay, MTT assay, GAG & DNA assay, RT-PCR, real time-PCR, and histological and immunochemical analysis in vitro and in vivo. Real-time PCR revealed that the expression of aggrecan and COMP by genetically modified hMSCs co-cultured with chondrocytes was 2 or 3 times greater than that of genetically modified MSCs alone. Moreover, the expression of collagen type II was more than 3.5 times greater than that of genetically modified MSCs alone. 3-D hydrogel constructs co-cultured with chondrocytes and genetically modified MSCs showed a significantly higher number of specific lacunae phenotypes at the end of the 4 week study, regardless of whether they were co-cultured in the presence of chondrocytes. These findings indicate that co-culture with chondrocytes and genetically modified MSCs can be used to engineer well designed implants for the formation of neocartilage by transplanted genetically modified MSCs.
Assuntos
Condrócitos/citologia , Condrogênese/fisiologia , Células-Tronco Mesenquimais/citologia , Animais , Sobrevivência Celular/genética , Sobrevivência Celular/fisiologia , Condrócitos/metabolismo , Condrogênese/genética , Citometria de Fluxo , Proteínas de Fluorescência Verde/genética , Proteínas de Fluorescência Verde/metabolismo , Hidrogel de Polietilenoglicol-Dimetacrilato , Imuno-Histoquímica , Células-Tronco Mesenquimais/metabolismo , Camundongos , Camundongos Nus , Coelhos , Reação em Cadeia da Polimerase Via Transcriptase ReversaRESUMO
In this work, a cocktail of dexamethasone (Dex) and transforming growth factor-beta3 (TGF-beta3) bound with heparin in/on PLGA microspheres was investigated by local dual delivery using rabbit mesenchymal stem cells (rMSCs) for chondrogenic differentiation in in vitro and in vivo study. In this study, we proved that the microsphere constructs were capable of simultaneously releasing TGF-beta3-conjugated with cy5.5 and Dex-conjugated with FITC with approximately zero order kinetics determined by Kodak imaging and confocal laser microscope. Microsphere constructs containing TGF-beta3 and Dex showed a significantly higher number of specific lacunae phenotypes at the end of the 4 week study irrespective of the presence of dexamethasone and TGF-beta3. Thus, dual delivery of TGF and Dex can be used to engineer inflammation-free and cartilage-associate tissue in the transplanted PLGA construct into nude mouse. These heparin-bound TGF-beta3-coated and Dex-loaded PLGA microsphere constructs show promise as coatings for implantable biomedical devices to improve biocompatibility and ensure both in vitro and in vivo performance.
Assuntos
Diferenciação Celular/efeitos dos fármacos , Dexametasona/farmacologia , Sistemas de Liberação de Medicamentos , Ácido Láctico/farmacologia , Células-Tronco Mesenquimais/citologia , Microesferas , Ácido Poliglicólico/farmacologia , Fator de Crescimento Transformador beta3/farmacologia , Animais , Proliferação de Células/efeitos dos fármacos , Forma Celular/efeitos dos fármacos , Fluoresceína-5-Isotiocianato/metabolismo , Perfilação da Expressão Gênica , Heparina/metabolismo , Hidrogel de Polietilenoglicol-Dimetacrilato , Imuno-Histoquímica , Células-Tronco Mesenquimais/efeitos dos fármacos , Células-Tronco Mesenquimais/metabolismo , Camundongos , Microscopia Confocal , Copolímero de Ácido Poliláctico e Ácido Poliglicólico , Ligação Proteica/efeitos dos fármacos , Coelhos , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Imagem Corporal TotalRESUMO
To encourage stem cell differentiation, gold nanoparticles (20 nm) were used to deliver electrical stimulation to human embryonic stem cells (hESCs) in vitro. Nano-structured gold nanoparticles were designed by coating the surface of culture dishes with gold nanoparticles using a layer-by-layer (LBL) system. In this method, gold nanoparticles were continuously coated onto dishes by SEM analysis. Evaluation of gene modified hESCs that were subsequently attached onto fibronectin-coated gold nanoparticles revealed that the un-differentiation marker, Oct-4, was no longer present following electrical stimulation. In addition, the osteogenic markers of collagen type I and Cbfa1 increased in response to electrical stimulation, while those of hESCs were not observed without electrical stimulation.