RESUMO
Adipose-derived stem cells (ASCs) can differentiate into multiple cell lineages and favor adipogenesis rather than osteogenesis. Because the extracellular matrix (ECM) component of the stem cell niche is important in stem cell differentiation, we hypothesized that ECM produced by human bone marrow stromal cells (BM-ECM) could enhance the osteogenic potential of ASCs during in vitro expansion. We have compared the replication and osteogenic differentiation of ASCs expanded on BM-ECM versus tissue culture plastic (TCP) in vitro and in vivo. During the first two passages, ASC proliferation on BM-ECM was 3.27-fold greater than that on TCP. ASCs expanded on BM-ECM formed more osteogenic colonies and higher expression of osteogenic markers than ASCs expanded on TCP. In nude mice, ASCs that had been expanded on BM-ECM formed more new bone tissue than those expanded on TCP. The data indicate that BM-ECM can be used to promote the osteogenic fate of ASCs.
Assuntos
Matriz Extracelular/química , Células-Tronco Mesenquimais/metabolismo , Células-Tronco/citologia , Tecido Adiposo/citologia , Animais , Células da Medula Óssea/citologia , Técnicas de Cultura de Células , Diferenciação Celular , Linhagem da Célula , Células Cultivadas , Subunidade alfa 1 de Fator de Ligação ao Core/metabolismo , Durapatita/química , Matriz Extracelular/metabolismo , Feminino , Células-Tronco Mesenquimais/citologia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Osteocalcina/metabolismo , Osteogênese , Transplante de Células-Tronco , Células-Tronco/metabolismoRESUMO
BACKGROUND: With goal of improving fat graft survival, many studies have focused on supplementing cells in the graft fat. In these studies, enhanced vascularization is considered the most important mechanism for the improved graft survival. Endothelial cells (ECs) are essential in vessel formation of the vascularization. Therefore, in this study, we coimplanted ECs with adipose tissue to investigate whether the ECs can enhance graft survival in a cell concentration-dependent manner. METHODS: Endothelial cells were isolated from stromal vascular fraction derived from human liposuction aspirates, and the EC characteristics were confirmed by CD31 immunofluorescence staining, measuring acetylated low-density lipoprotein uptake, and observing the formation of capillary-like tubular structures in Matrigel. During the animal experiment, the isolated ECs were labeled, then added to 0.5-mL fat grafts at different numbers (0.5 × 10(6), 1 × 10(6), 2 × 10(6), and 4 × 10(6) cells) before subcutaneous implantation in nude mice. Grafts were harvested at 1 week, 1 month, and 2 months after -transplantation, and graft survival and vascularization were evaluated based on weight measurements, histological assessment, and vascular gene expression. RESULTS: Stromal vascular fraction-derived vascular cells exhibited typical EC characteristics. The observed differences in explanted graft weight, vessel density, vascular gene expression, and cell tracking result indicated that coimplantation with ECs accelerated vascularization that increased graft survival in a concentration-dependent manner. Over the experimental period, fat grafts implanted with 4 × 10(6) ECs showed no weight loss and the greatest increases in measures of vascularization. CONCLUSIONS: Endothelial cells can effectively enhance vascularization in fat grafts, and higher EC concentrations (eg, 4 × 10(6) ECs/0.5 mL adipose tissue) may best support graft survival.
Assuntos
Tecido Adiposo/transplante , Endotélio Vascular/transplante , Sobrevivência de Enxerto/fisiologia , Neovascularização Fisiológica/fisiologia , Lesões dos Tecidos Moles/cirurgia , Animais , Células Cultivadas , Modelos Animais de Doenças , Endotélio Vascular/citologia , Feminino , Humanos , Camundongos , Camundongos Nus , Lesões dos Tecidos Moles/patologiaRESUMO
The effect of focal adhesion kinase (FAK) on suppressing scarring and the potential molecular mechanism underlying it has been investigated. Ten samples of human hypertrophic scars (HS) tissue cultured in vitro were transfected with FAK siRNA mediated by liposome. Quantitative real-time PCR was used to detect the expression of integrin α, transforming growth factor-ß (TGF-ß), FAK and α-smooth muscle actin (α-SMA) after transfection. MTT assay was used as a measure of fibroblast proliferation. Flow cytometry and (3)H-proincorporation technique gave measurements of the cell cycle and the quantity of collagen synthesis, respectively. Expression of FAK was effectively blocked, accompanied by decreasing expression of integrin α, TGF-ß and α-SMA in hypertrophic scars fibroblast (HSFB) cells. One to 4 h after transfection with FAK siRNA, proliferation of HSFB cells was strongly inhibited (P < 0.01), reaching a maximum at 48 h. The proportion of G1 cells was higher and the proportion of the S and G2 cells lower after transfection. The amount of collagen synthesis in HSFB cells decreased when HSFB cells were transfected for 48 h. RNA interference targeting the FAK gene can block the two abnormal signal transduction pathways mediated by the integrin and TGF-ß receptors that are responsible for hyperplasia and contracture of the scar, making FAK iRNA therapy a potentially effective approach in HS treatment.
Assuntos
Actinas/metabolismo , Cicatriz Hipertrófica/metabolismo , Quinase 1 de Adesão Focal/metabolismo , Cadeias alfa de Integrinas/metabolismo , RNA Interferente Pequeno/metabolismo , Fator de Crescimento Transformador beta/metabolismo , Actinas/genética , Adulto , Células Cultivadas , Cicatriz Hipertrófica/patologia , Colágeno/biossíntese , Quinase 1 de Adesão Focal/antagonistas & inibidores , Quinase 1 de Adesão Focal/genética , Pontos de Checagem da Fase G1 do Ciclo Celular , Humanos , Cadeias alfa de Integrinas/genética , Interferência de RNA , RNA Mensageiro/metabolismo , Receptores de Fatores de Crescimento Transformadores beta/metabolismo , Fatores de Tempo , Fator de Crescimento Transformador beta/genética , Adulto JovemRESUMO
OBJECTIVES: In order to explore the molecular mechanism of hydrocortisone in keloid tissue, the gene expression profiles of keloid samples treated with hydrocortisone were subjected to bioinformatics analysis. METHODS: Firstly, the gene expression profiles (GSE7890) of five samples of keloid treated with hydrocortisone and five untreated keloid samples were downloaded from the Gene Expression Omnibus (GEO) database. Secondly, data were preprocessed using packages in R language and differentially expressed genes (DEGs) were screened using a significance analysis of microarrays (SAM) protocol. Thirdly, the DEGs were subjected to gene ontology (GO) function and KEGG pathway enrichment analysis. Finally, the interactions of DEGs in samples of keloid treated with hydrocortisone were explored in a human protein-protein interaction (PPI) network, and sub-modules of the DEGs interaction network were analyzed using Cytoscape software. RESULTS: Based on the analysis, 572 DEGs in the hydrocortisone-treated samples were screened; most of these were involved in the signal transduction and cell cycle. Furthermore, three critical genes in the module, including COL1A1, NID1, and PRELP, were screened in the PPI network analysis. CONCLUSIONS: These findings enhance understanding of the pathogenesis of the keloid and provide references for keloid therapy.
Assuntos
Regulação da Expressão Gênica , Hidrocortisona/uso terapêutico , Queloide/tratamento farmacológico , Queloide/genética , Mapas de Interação de Proteínas , HumanosRESUMO
Impressive results in patient care and cost reduction have increased the demand for systems-engineering methodologies in large health care systems. This Report from the Field describes the feasibility of applying systems-engineering techniques at a community health center currently lacking the dedicated expertise and resources to perform these activities.
Assuntos
Centros Comunitários de Saúde/organização & administração , Engenharia/métodos , Eficiência Organizacional , Prática Clínica Baseada em Evidências/métodos , Humanos , Modelos Organizacionais , Melhoria de Qualidade/organização & administração , Estudos de Tempo e MovimentoRESUMO
OBJECTIVE: To evaluate the clinical effect of cell-assisted lipotransfer (CAL) for breast augmentation. METHODS: 18 patients accepted breast augmentation using CAL. 10 patients completed 6-month follow-up and were involved in the study. The adipose tissue was harvested from patients' thighs, flanks and lower abdomen with Lipokit. After standing, 250 ml fatty portion and 500 ml fluid portion of suction aspirates were processed according to the procedures reported in reference. Flow-cytometry was used to detect the percentage of adipose-derived stem cells(ADSCs) in distilled stromal vascular fraction (SVF). The differentiation function of cultured cells also was assessed. The breast volume and images were evaluated by using MRI before operation, 3 and 6 months after operation. The breast volume was marked as V0, V3 and V6 respectively. The resorption rate of transplanted adipose tissue for each breast was calculated and marked as R3 and R6. RESULTS: Averagely, the percentage of ADSCs in freshly distilled SVF was 41.67%. The in-vitro cultured cell grew well and could differentiate into fat, bone and cartilage. Statistics showed that V0, V3 and V6 was (416.19 +/- 40.43) ml, (551.72 +/- 59.86) ml and (538.81 +/- 68.35) ml respectively. R3 and R6 was (51.20 +/- 11.96)% and (54.22 +/- 12.73)%. There was significant difference between V3 and V0 (P < 0.05), V6 and V0. However, no significant difference was showed between V3 and V6 or R3 and R6. In addition, no cyst or calcification was seen in all MRI images. CONCLUSIONS: In process of breast augmentation using CAL, the distilled SVF contains 41.67% ADSCs which have adipogenic, osteogenic and chondrogenic function. Within 3-month post-operation, the breast volume decreases obviously but the volume sustains after that. Compared with the preoperative volume, the 6-month postoperative volume is significantly increased and the breasts' contour is improved greatly. This study indicates that CAL is a safe and effective way for breast augmentation.
Assuntos
Adipócitos/transplante , Mamoplastia/métodos , Transplante de Células-Tronco , Adipócitos/citologia , Tecido Adiposo/citologia , Adulto , Diferenciação Celular , Células Cultivadas , Feminino , Humanos , Pessoa de Meia-Idade , Células Estromais/citologiaRESUMO
OBJECTIVE: To study the role of focal adhesion kinase (FAK) in the pathogenesis of human hypertrophic scar. METHODS: Human hypertrophic scar fibroblasts (HSFB) were isolated from human hypertrophic scar and cultured in vitro. The cells were then divided into 3 groups as AT group (phosphorothioate FAK ASODN was transfected into the HSFB by liposome), LPC group (liposome only), and LC group (control group, without liposome or ASODN). The FAKmRNA index of HSFB was assessed by polymerase chain reaction method (FQ-PCR). The collagen synthesis of HSFB was assessed by 3H-proline incorporation method. RESULTS: The FAK mRNA index of HSFB in AT group 48 hours after transfection was significantly lower than that in LPC and LC groups (0.043 +/- 0.030, 0.124 +/- 0.070, 0.127 +/- 0.0195, P < 0.05). The 3H-proline incorporation rate in AT group was lower than that in LPC and LC groups (257.0 +/- 15.14, 962.2 +/- 300.5, 930.8 +/- 28.97, P < 0.01). CONCLUSION: The expression of FAK gene and collagen synthesis of the cultured HSFB could be inhibited by FAK ASODN, indicating that FAK played a role in the development of excessive fibrosis of human hypertrophic scar.