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1.
Adv Exp Med Biol ; 1131: 1065-1078, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-31646545

RESUMO

Our blood serum Ca2+ levels are maintained within a narrow range (Ca2+ homeostasis) through a complex feedback system. However, local bone marrow Ca2+ levels can reach high concentrations, at least transiently, due to bone resorption, which is one of the notable features of the bone marrow stroma. Bone homeostasis is maintained by both the balance between osteoblastic bone formation and osteoclastic bone resorption and the balance of mesenchymal stem cell differentiation into osteoblasts and adipocytes. It has been reported that under culture conditions of infrequent adipocyte differentiation (no treatment with insulin or dexamethasone), high extracellular Ca2+ enhances osteoblast but not adipocyte accumulation in bone marrow stromal cells. In contrast, under culture conditions of predominant adipocyte differentiation (treatment with insulin and dexamethasone), high extracellular Ca2+ enhances adipocyte but not osteoblast accumulation in bone marrow stromal cells. Thus, the increased extracellular Ca2+ caused by bone resorption might enhance osteoblast development to reform missing bone under conditions of infrequent adipocyte differentiation (such as the normal physiological state) and might accelerate adipocyte accumulation instead of osteoblastic bone formation under conditions of predominant adipocyte differentiation (such as aging, obesity, use of glucocorticoids, and postmenopause). Moreover, increased adipocyte accumulation in bone marrow suppresses lymphohematopoiesis and contributes to a dysfunction of osteogenesis.


Assuntos
Medula Óssea , Cálcio , Células-Tronco Mesenquimais , Adipócitos/citologia , Medula Óssea/metabolismo , Células da Medula Óssea/citologia , Células da Medula Óssea/patologia , Cálcio/metabolismo , Diferenciação Celular , Humanos , Células-Tronco Mesenquimais/citologia , Osteoblastos/citologia
2.
Zhonghua Shao Shang Za Zhi ; 35(9): 641-644, 2019 Sep 20.
Artigo em Chinês | MEDLINE | ID: mdl-31594181

RESUMO

Adipose stem cells (ASCs) are mesenchymal stem cells derived from adipose tissue, and they have potentials of self-renewal and multi-directional differentiation. Compared with bone marrow mesenchymal stem cells, ASCs have many advantages, such as easy access, easy cultivation, and abundant content, which are valuable seed cells in the field of repair and reconstruction. In recent years, with the deepening of the researches on differentiation, regulation, and function of ASCs, the clinical application of ASCs has gradually increased with good therapeutic effects.


Assuntos
Adipócitos/citologia , Terapia Baseada em Transplante de Células e Tecidos , Células-Tronco Mesenquimais/citologia , Engenharia Tecidual , Tecido Adiposo/citologia , Diferenciação Celular , Humanos
3.
Int J Nanomedicine ; 14: 7795-7808, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31576124

RESUMO

Background: Endogenously expressed microRNAs (miRNAs) have attracted attention as important regulators in post-transcriptionally controlling gene expression of various physiological processes. As miRNA dysregulation is often associated with various disease patterns, such as obesity, miRNA-27a might therefore be a promising candidate for miRNA mimic replacement therapy by inhibiting adipogenic marker genes. However, application of naked nucleic acids faces some limitations concerning poor enzymatic stability, bio-membrane permeation and cellular uptake. To overcome these obstacles, the development of appropriate drug delivery systems (DDS) for miRNAs is of paramount importance. Methods: In this work, a triple combination of atomic force microscopy (AFM), brightfield (BF) and fluorescence microscopy was used to trace the cellular adhesion of N-TER peptide-nucleic acid complexes followed by time-dependent uptake studies using confocal laser scanning microscopy (cLSM). To reveal the biological effect of miRNA-27a on adipocyte development after transfection treatment, Oil-Red-O (ORO)- staining was performed to estimate the degree of in lipid droplets accumulated ORO in mature adipocytes by using light microscopy images as well as absorbance measurements. Results: The present findings demonstrated that amphipathic N-TER peptides represent a suitable DDS for miRNAs by promoting non-covalent complexation through electrostatic interactions between both components as well as cellular adhesion of the N-TER peptide - nucleic acid complexes followed by uptake across cell membranes and intracellular release of miRNAs. The anti-adipogenic effect of miRNA-27a in 3T3-L1 cells could be detected in mature adipocytes by reduced lipid droplet formation. Conclusion: The present DDS assembled from amphipathic N-TER peptides and miRNAs is capable of inducing the anti-adipogenic effect of miRNA-27a by reducing lipid droplet accumulation in mature adipocytes. With respect to miRNA mimic replacement therapies, this approach might provide new therapeutic strategies to prevent or treat obesity and obesity-related disorders.


Assuntos
Sistemas de Liberação de Medicamentos , MicroRNAs/metabolismo , Peptídeos/química , Células 3T3-L1 , Adipócitos/citologia , Adipócitos/metabolismo , Adipogenia , Sequência de Aminoácidos , Animais , Adesão Celular , Gotículas Lipídicas/metabolismo , Camundongos , MicroRNAs/genética , Ácidos Nucleicos Peptídicos/química , Transfecção
4.
Res Vet Sci ; 125: 351-359, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31374445

RESUMO

Fibronectin type III domain-containing protein 5 (FNDC5) plays an important role in white-to-brown adipose tissue conversion in mice. However, there is no report on the role of this protein in Arbas Cashmere goat adipose tissue. We investigated the effect of FNDC5 on the proliferation and differentiation of goat adipose-derived stem cells (gADSCs). We found that FNDC5 promotes the proliferation of gADSCs and increases the levels of p-ERK and p-p38, while it has no effect on the levels of ERK, p38, AKT and p-AKT. What's more, FNDC5 promotes the differentiation of gADSCs into lipid droplets. Overexpression of FNDC5 increased the protein levels of ASC1, UCP1, PPARγ and SREBP1. Additionally, mRNA levels of PPARγ, SREBP1, ACC, and FABP4 increased significantly. FNDC5 knockdown was associated with opposite effects. These results suggest that FNDC5 promotes the proliferation of gADSCs via MAPK signaling pathway and also induces the differentiation of these cells.


Assuntos
Adipócitos/efeitos dos fármacos , Diferenciação Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Fibronectinas/farmacologia , Células-Tronco Mesenquimais/efeitos dos fármacos , Adipócitos/citologia , Animais , Células-Tronco Mesenquimais/citologia , Camundongos
5.
DNA Cell Biol ; 38(9): 945-954, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31355674

RESUMO

Domestic cattle are an important type of livestock, with beef production playing a major role in the agricultural economy. Adipocyte levels and fat content are interrelated, with meat quality being highly dependent on its fat content and distribution. Acyl-CoA synthetases of long-chain (ACSL) fatty acids (FAs) play an integral role in virtually every metabolic pathway in mammalian biochemistry, including complex lipid biosynthesis, protein modification, and ß-oxidation processes. ACSL3 activity is also known to be associated with adipocyte differentiation; however, its biological mechanism of action is currently unclear. Gene expression in subcutaneous preadipocytes isolated from subcutaneous deposits of Chinese Red Steppe cattle has been studied using in vitro cell transfection, real-time polymerase chain reaction and western blot analysis. The lipid and triglyceride contents of lipid droplets have also been measured to verify the levels of gene expression. These combined studies show that ACSL3 is induced during adipocyte differentiation, with its overexpression promoting an increase in the triglyceride content of lipid droplets. Furthermore, mRNA and protein expression levels for adipocyte differentiation marker genes, such as peroxisome proliferator-activated receptor gamma (PPARγ) and CCAAT/enhancer-binding protein alpha (C/EBPα), were markedly increased during mature adipocyte cell differentiation. Knockdown of ACSL3 expression using ACSL3 small interfering RNAs (siRNAs) resulted in a decrease in lipid content of cattle adipocytes, providing further evidence that ACSL3 plays a key role in the differentiation process.


Assuntos
Adipócitos/citologia , Adipogenia , Bovinos/genética , Coenzima A Ligases/genética , Adipócitos/metabolismo , Animais , Proteína alfa Estimuladora de Ligação a CCAAT/genética , Proteína alfa Estimuladora de Ligação a CCAAT/metabolismo , Células Cultivadas , Coenzima A Ligases/metabolismo , PPAR gama/genética , PPAR gama/metabolismo
6.
Plast Reconstr Surg ; 144(2): 207e-217e, 2019 08.
Artigo em Inglês | MEDLINE | ID: mdl-31348343

RESUMO

BACKGROUND: Assisted lipotransfer for breast reconstruction involves the isolation and supplementation of adipose-derived stromal cells. This procedure has raised concerns regarding safety with respect to promotion of tumor growth and relapse. Several in vitro and animal experimental studies have indicated increased survival, growth, and invasive characteristics of breast cancer cells on interaction with adipose-derived stromal cells. These results seem to be in poor concordance with clinical observations of a low rate of cancer recurrences after assisted lipotransfer. METHODS: The authors investigated the effects of adipose-derived stromal cells and adipose-derived stromal cells differentiated into adipocytes and fibroblasts on five breast cancer cell lines (i.e., T47D, MCF-7, BT20, MDA-MB-231, and ZR-75-1) and MCF-10A, a nonmalignant counterpart. RESULTS: Conditioned media of adipose-derived stromal cells stimulated the proliferation of breast cancer cell lines depending on the individual adipose-derived stromal cell-breast cancer cell line combination. Conditioned media of adipose-derived stromal cells differentiated into adipocytes gave a lower response, and conditioned media of fibroblasts were also active. A putative cancer stem cell-like phenotype was not increased by adipose-derived stromal cell-conditioned media, no physical interaction of cancer cells with adipose-derived stromal cells was detectable on scanning electron microscopy, and cell migration was not enhanced. Adipogenic differentiation of adipose-derived stromal cells indicated that hepatocyte growth factor, insulin-like growth factor-binding protein-3, insulin-like growth factor-binding protein-6, interleukin-6, CCL2/MCP-1, and macrophage colony-stimulating factor are not linked to the proliferative activity of conditioned media. CONCLUSION: The results indicate that the adipose-derived stromal cells used for assisted lipotransfer are not expected to increase the risk of tumor recurrence to a major degree in correspondence with the clinical observation of the affected breast cancer patients. CLINICAL QUESTION/LEVEL OF EVIDENCE: Therapeutic, V.


Assuntos
Adipócitos/citologia , Comunicação Celular/fisiologia , Diferenciação Celular/fisiologia , Western Blotting , Neoplasias da Mama/patologia , Neoplasias da Mama/cirurgia , Estudos de Casos e Controles , Linhagem Celular Tumoral , Movimento Celular/fisiologia , Proliferação de Células/fisiologia , Técnicas de Cocultura , Feminino , Humanos , Valores de Referência , Sensibilidade e Especificidade , Células Estromais/citologia
7.
J Agric Food Chem ; 67(32): 8839-8846, 2019 Aug 14.
Artigo em Inglês | MEDLINE | ID: mdl-31334651

RESUMO

Natural products are one of the main sources for discovering new lead compounds. We previously reported that cinnamon extract has a promising effect in regulating lipid tissue volume and insulin sensitivity in vivo. However, its effective component and the underlying mechanism are not known. In the present study, we analyzed the effect of different components of cinnamon on regulating insulin sensitivity in 3T3-L1 adipocytes. Functional assay revealed that, of the six major components of cinnamon extracts, the B-type procyanidin, procyanidin C1, improves the differentiation of 3T3-L1 cells (TG content: 1.10 ± 0.09 mM at a dosage of 25 µM vs 0.67 ± 0.02 mM in vehicle group, p < 0.001) and promotes insulin-induced glucose uptake (8.58 ± 1.43 at a dosage of 25 µM vs 3.05 ± 1.24 in vehicle group, p < 0.001). Mechanism studies further suggested that procyanidin C1 activates the AKT-eNOS pathway, thus up-regulating glucose uptake and enhancing insulin sensitivity in mature adipocytes. Taken together, our study identified B-type procyanidin C1, a component of cinnamon extract, that stimulates preadipocyte differentiation and acts as a potential insulin action enhancer through the AKT-eNOS pathway in mature adipocytes.


Assuntos
Adipócitos/efeitos dos fármacos , Adipócitos/metabolismo , Biflavonoides/farmacologia , Catequina/farmacologia , Cinnamomum zeylanicum/química , Insulina/metabolismo , Extratos Vegetais/farmacologia , Proantocianidinas/farmacologia , Células 3T3-L1 , Adipócitos/citologia , Animais , Transporte Biológico/efeitos dos fármacos , Diferenciação Celular/efeitos dos fármacos , Glucose/metabolismo , Camundongos , Óxido Nítrico Sintase Tipo III/genética , Óxido Nítrico Sintase Tipo III/metabolismo , Proteínas Proto-Oncogênicas c-akt/genética , Proteínas Proto-Oncogênicas c-akt/metabolismo
8.
Cytogenet Genome Res ; 158(3): 133-144, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31272101

RESUMO

Bone morphogenetic protein 2 (BMP2) can mediate the signaling of R-Smads and regulate different biological functions, including adipocyte differentiation. Long noncoding RNAs (lncRNAs) can be involved in many important biological processes, including fat metabolism, as miRNA sponges. This study aimed to investigate the molecular mechanism of fat deposition and to provide useful information for the prevention and treatment of lipid-related diseases. lncRNA sequencing was performed to compare and analyze, for the first time, the expression of lncRNAs in BMP2-induced and non-BMP2-induced preadipocytes from Junmu1 pigs. In addition, functional annotation and enrichment analysis of differentially expressed lncRNA target genes were carried out. lncRNAs and mRNAs were compared and analyzed. lncRNAs were identified that may regulate adipogenesis and lipid metabolism. The results give a theoretical basis for further studies on fat deposition mechanisms and provide potential therapeutic targets for metabolic diseases.


Assuntos
Adipócitos/efeitos dos fármacos , Proteína Morfogenética Óssea 2/farmacologia , Diferenciação Celular/efeitos dos fármacos , RNA Longo não Codificante/análise , Células-Tronco/efeitos dos fármacos , Suínos/genética , Transcriptoma/genética , Adipócitos/citologia , Adipócitos/metabolismo , Animais , RNA Longo não Codificante/genética , Análise de Sequência de RNA , Células-Tronco/citologia , Células-Tronco/metabolismo , Triglicerídeos/metabolismo
9.
Zhonghua Gan Zang Bing Za Zhi ; 27(6): 450-456, 2019 Jun 20.
Artigo em Chinês | MEDLINE | ID: mdl-31357762

RESUMO

Objective: To observe the effect of differentiated mature adipocytes on hepatic steatosis and aquaporin-9 (AQP9) expressions in HepG2 cells and further explore its possible mechanism of action. Methods: Human preadipocytes were cultured and differentiated to full maturity. HepG2 cells were co-cultured with non-differentiated adipocytes and differentiated mature adipocytes for 48 h, and then labeled as control group and experimental group. Oil red O staining and intracellular triglyceride content were performed on co-cultured HepG2 cells and simultaneous changes in phosphatidylinositol 3-kinase (PI3K) - serine/threonine kinase (Akt) signaling pathway, and AQP9 mRNA and protein levels were detected. The experimental group was co-cultured with recombinant human insulin-like growth factor-I (IGF-I), with the addition of 100ng/ml PI3K-Akt pathway agonist, labeled as experimental group + IGF-I group. The activation of PI3K-Akt pathway was verified by Western blotting (WB). The expression of AQP9 was detected by RT-q PCR and WB. The recombinant lentivirus LV-AQP9 or empty-loaded virus LV-PWPI was transfected with HepG2 cells by recombinant lentiviral transfection tecnique, and labeled as HepG2-AQP9 and HepG2-PWPI. The transfection efficiency was assessed by confocal laser scanning microscopy and RT-qPCR and WB detected the change of AQP9 expression level after virus transfection. Afterwards, the stable over-expressed HepG2-AQP9 cells and the empty-loaded HepG2-PWPI cells were co-cultured with differentiated mature adipocytes for 48h, and labeled as HepG2-AQP9 co-culture group, and then intracellular triglyceride content were detected with Oil red O staining. Finally, IGF-I was added to the HepG2-AQP9 co-culture group, which was recorded as HepG2-AQP9 co-culture + IGF-I group. Intracellular triglyceride content was detected with Oil red O staining, and WB verified PI3K-Akt signaling pathway activation and changes in AQP9 mRNA and protein levels. A t-test was used to compare the two independent samples. Results: The intracellular lipid droplets and triglyceride content (0.052 ± 0.005) in the experimental group was increased significantly than the control group (0.033 ± 0.003) (t= 5.225,P= 0.006), suggesting that adipocyte co-culture had induced steatosis in HepG2 cells. RT-qPCR and WB results indicated that the expression levels of AQP9 mRNA (3.615 ± 0.330) and protein levels (0.072 ± 0.005) in the experimental group were significantly higher than the control group (t= 13.708, 11.225,P= 0.005, < 0.001). WB results showed that the expression level of phosphorylated Akt (p-Akt) protein (0.116±0.003) in the experimental group was significantly lower than the control group (0.202 ± 0.003) (t= 27.136,P< 0.001). The total Akt protein was constant, and the p-Akt/total Akt (0.182 ± 0.017)was significantly lower than the control group (0.327 ± 0.019) (t= 2.431,P= 0.001), suggesting that adipocyte co-culture had inhibited PI3K- Akt signaling pathway in HepG2 cells and up-regulated the expression level of AQP9. WB results indicated that the expression level of p-Akt protein (0.194 ± 0.021) in the experimental group + IGF-I group was significantly higher than the experimental group (0.132 ± 0.003) (t= 5.082,P= 0.007). The total Akt protein was constant, and the p-Akt/total Akt (0.281 ± 0.009) was significantly higher than the control group (0.184 ± 0.132) (t= 10.311,P< 0.001). Simultaneously, RT-qPCR and WB results indicated that the expression levels of AQP9 mRNA (0.327 ± 0.347) and protein levels (0.042 ± 0.004) in the experimental group + IGF-I group were significantly lower than the experimental group (t= 33.573, 5.598,P< 0.001, 0.005), suggesting that adipocyte co-culture had possibility to regulate the expression level of AQP9 through the PI3K-Akt pathway. Confocal laser microscopy analysis showed that the transfection efficiency was more than 90%. RT-q PCR and WB results indicated that the expression levels of AQP9 mRNA and protein levels (0.373 ± 0.221) in HepG2-AQP9 group were significantly higher than HepG2-PWPI group (t=14.953, 28.931,P= 0.002 and 0.000), suggesting that the stable overexpression of AQP9 cell line was successfully constructed. The intracellular lipid droplets and triglyceride content in HepG2-AQP9 co-culture group was significantly increased (t= 5.478, 5.369,P= 0.005) than HepG2-PWPI co-culture group and HepG2-AQP9 co-culture+ IGF-I group, suggesting that the increased expression of AQP9 had promoted HepG2 steatosis in co-cultured adipocytes. WB results showed the expression levels of p-Akt protein (0.168 ± 0.006) and p-Akt/total Akt (0.265±0.009) in HepG2-AQP9 co-culture + IGF-1 group was significantly increased (t= 16.311, 8.769,P< 0.001) than HepG2-AQP9 co-culture group, while the expression levels of AQP9 mRNA (0.327 ± 0.034) and protein (0.375 ± 0.025) was significantly decreased (t= 33.573, 9.146,P< 0.001 and 0.001). Conclusion: Adipocytes co-culture can induce steatosis in HepG2 cells, and may participate in inhibiting PI3K-Akt signaling pathway to upregulate the expression of AQP9 in steatotic HepG2 cells.


Assuntos
Adipócitos , Aquaporinas , Regulação da Expressão Gênica no Desenvolvimento , Adipócitos/citologia , Adipócitos/metabolismo , Aquaporinas/genética , Técnicas de Cocultura , Células Hep G2 , Humanos , Fosfatidilinositol 3-Quinases/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo
10.
Int J Mol Sci ; 20(11)2019 Jun 05.
Artigo em Inglês | MEDLINE | ID: mdl-31195622

RESUMO

Diabetes is a contributor to morbidity across the globe and is often associated with obesity, metabolic syndrome and other inflammatory diseases associated with aging. In addition to genetic and lifestyle factors, environmental factors such as metals and persistent organic pollutants may increase the severity or lower the threshold of these conditions. In cell culture, methylmercury is toxic to adipocytes and may impact adipokine secretions. In this study, we determined the effects of different concentrations of theaflavin digallate on methylmercury exposed 3T3-L1 adipocytes in cell culture. Secretions of resistin, adiponectin and lipid peroxidation product, 4-hydroxynonenal (4-HNE) were monitored using ELISA assays. Cell morphology of methylmercury and theaflavin-3,3'-digallate treated adipocytes was assessed using Lipid (Oil Red O) staining. Exposure to methylmercury increased the levels of resistin and adiponectin as well as 4-HNE when compared to the control cells. Methylmercury treated cells resulted in smaller number of adipocytes and clumped lipid droplets. These results suggest that methylmercury induces reactive oxygen species leading to development of an inflammatory response. Theaflavin-3,3'-digallate reduced the impact of methylmercury by maintaining the adipocytes morphology and secretion patterns of adiponectin, resistin and 4-hydroxynonenal. With this experimental model system other anti-inflammatory and signaling agents could be tested at the biochemical level before eventually leading to studies in animal models.


Assuntos
Adipócitos/citologia , Adipocinas/metabolismo , Biflavonoides/farmacologia , Catequina/farmacologia , Diferenciação Celular , Ácido Gálico/análogos & derivados , Compostos de Metilmercúrio/toxicidade , Células 3T3-L1 , Adipócitos/efeitos dos fármacos , Adipócitos/metabolismo , Adiponectina/metabolismo , Aldeídos/metabolismo , Animais , Diferenciação Celular/efeitos dos fármacos , Forma Celular/efeitos dos fármacos , Ácido Gálico/farmacologia , Camundongos , Resistina/metabolismo
11.
Biochemistry (Mosc) ; 84(5): 553-561, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-31234769

RESUMO

Obesity is accompanied by dyslipidemia, hypoxia, endoplasmic reticulum (ER) stress, and inflammation, representing the major risk factor for the development of insulin resistance (IR) and type 2 diabetes. We modeled these conditions in cultured 3T3-L1 adipocytes and studied their effect on insulin signaling, glucose uptake, and inflammatory response via activation of stress-dependent JNK1/2 kinases. Decreased insulin-induced phosphorylation of the insulin cascade components IRS, Akt, and AS160 was observed under all tested conditions (lipid overloading of cells by palmitate, acute inflammation induced by bacterial lipopolysaccharide, hypoxia induced by Co2+, and ER stress induced by brefeldin A). In all the cases, except the acute inflammation, glucose uptake by adipocytes was reduced, and the kinetics of JNK1/2 activation was bi-phasic exhibiting sustained activation for 24 h. By contrast, in acute inflammation, JNK1/2 phosphorylation increased transiently and returned to the basal level within 2-3 h of stimulation. These results suggest a critical role of sustained (latent) vs. transient (acute) inflammation in the induction of IR and impairment of glucose utilization by adipose tissue. The components of the inflammatory signaling can be promising targets in the development of new therapeutic approaches for preventing IR and type 2 diabetes.


Assuntos
Inflamação , Resistência à Insulina , Obesidade/patologia , Células 3T3-L1 , Adipócitos/citologia , Adipócitos/efeitos dos fármacos , Adipócitos/metabolismo , Animais , Estresse do Retículo Endoplasmático/efeitos dos fármacos , Ácidos Graxos não Esterificados/farmacologia , Inflamação/etiologia , Insulina/farmacologia , Lipopolissacarídeos/farmacologia , Camundongos , Proteína Quinase 8 Ativada por Mitógeno/metabolismo , Proteína Quinase 9 Ativada por Mitógeno/metabolismo , Obesidade/metabolismo , Fosfatidilinositol 3-Quinases/metabolismo , Fosforilação/efeitos dos fármacos , Proteínas Proto-Oncogênicas c-akt/metabolismo , Transdução de Sinais/efeitos dos fármacos
12.
Int J Mol Sci ; 20(11)2019 May 29.
Artigo em Inglês | MEDLINE | ID: mdl-31146351

RESUMO

Evaluating cell migration after cell-based treatment is important for several disorders, including osteoarthritis (OA), as it might influence the clinical outcome. This research explores migrating expanded-adipose stromal cells (ASCs) and adipose niches after enzymatic and mechanical processes. Bilateral anterior cruciate ligament transection induced a mild grade of OA at eight weeks in adult male New Zealand rabbits. ASCs, enzymatic stromal vascular fraction (SVF), and micro fragmented adipose tissue (MFAT) were intra-articularly injected in the knee joint. Assessments of cell viability and expression of specific markers, including CD-163 wound-healing macrophages, were done. Cell migration was explored through labelling with PKH26 dye at 7 and 30 days alongside co-localization analyses for CD-146. All cells showed good viability and high percentages of CD-90 and CD-146. CD-163 was significantly higher in MFAT compared to SVF. Distinct migratory potential and time-dependent effects were observed among cell-based treatments. At day 7, both ASCs and SVF migrated towards synovium, whereas for MFAT versus cartilage, a different migration pattern was noticed at day 30. The long-term distinct cell migration of ASCs, SVF, and MFAT open interesting clinical insights on their potential use for OA treatment. Moreover, the highest expression of CD-163 in MFAT, rather than SVF, might have an important role in directly mediating cartilage tissue repair responses.


Assuntos
Adipócitos/transplante , Osteoartrite do Joelho/terapia , Regeneração , Transplante de Células-Tronco/métodos , Adipócitos/citologia , Adipócitos/fisiologia , Animais , Antígenos de Diferenciação/genética , Antígenos de Diferenciação/metabolismo , Movimento Celular , Células Cultivadas , Condrócitos/citologia , Condrócitos/metabolismo , Macrófagos/citologia , Macrófagos/metabolismo , Masculino , Cultura Primária de Células/métodos , Coelhos
13.
Mol Biol (Mosk) ; 53(3): 497-501, 2019.
Artigo em Russo | MEDLINE | ID: mdl-31184615

RESUMO

Homeodomain transcription factors play a significant role in adipocyte differentiation. The role of Pbx1 and Prep1, proteins of the TALE family (the three amino acid loop extension), was previously established in adipocyte differentiation of mesenchymal stromal cells and 3T3-L1 cell line. In this study, with the use of RNA interference technology we show that another transcription factor from the same family, Meis1, which is a core protein of mature cardiomyocytes, represses adipogenesis to a greater degree than its paralog Meis2. A number of Meis target genes, markers of adipocytes, are identified. This may indicate the transcriptional mechanism of the effect of Meis1 on the adipocyte differentiation of mouse preadipocytes.


Assuntos
Adipócitos/citologia , Diferenciação Celular , Proteína Meis1/metabolismo , Miócitos Cardíacos/metabolismo , Adipócitos/metabolismo , Animais , Diferenciação Celular/genética , Proteínas de Homeodomínio/metabolismo , Camundongos , Células-Tronco/citologia , Células-Tronco/metabolismo
14.
Gene ; 710: 406-414, 2019 Aug 20.
Artigo em Inglês | MEDLINE | ID: mdl-31200087

RESUMO

Integrins are cell attachment receptors that function in the communication between the intracellular and extracellular compartments. Integrins and extra cellular matrix (ECM) collaborate to regulate gene expression by extracellular signal-regulated kinases (ERKs). Integrins as regulators, have critical role in ECM remodeling. Fibrosis is the hallmark of obesity and insulin resistance originated by aberrant ECM remodeling. Therefore deciphering integrins' expression profile in cells at different conditions is worthy. The aim of this study is evaluation of integrins' gene expression profile changes in mouse 3T3-L1 preadipocytes, adipocytes, insulin resistant and hypertrophic adipocytes. For this purpose, we differentiated mouse 3T3-L1 preadipocytes to adipocytes, insulin resistant and hypertrophied adipocytes and assayed integrins' gene expression in four conditions by real time-PCR. Also the proteins expression changes of ERK and collagen VI assayed by Western blotting. Data analysis has shown that integrins' gene expression changes throughout adipocyte differentiation and pathological processes. The expressions of many integrins genes were significantly up- or down-regulated by >1.5-fold during differentiation, insulin resistant, and hypertrophic adipocytes. In addition to changes in the type of integrin, the integrins expression levels were different. Integrins, on the whole were more expressed in pathological processes relative to normal adipocytes. Also, phosphorylation of ERK 1,2 was increased >1.5-fold in differentiated, insulin resistant and hypertrophied adipocytes versus preadipocytes. Collagen VI only increased 2-fold in hypertrophied adipocytes. Examination of the total integrin gene family expression during adipocyte differentiation and pathological processes, leads to the identification of differential integrin gene expression. These results suggest that the type of integrin may not only play a role in adipocyte differentiation but also in pathological processes which may associate to increased ERK pathway activity in these conditions.


Assuntos
Adipócitos/citologia , Adipócitos/patologia , Perfilação da Expressão Gênica/métodos , Integrinas/genética , Células 3T3-L1 , Adipócitos/metabolismo , Animais , Diferenciação Celular , Colágeno Tipo VII/metabolismo , Regulação da Expressão Gênica , Hipertrofia , Resistência à Insulina , Sistema de Sinalização das MAP Quinases , Camundongos , Família Multigênica , Fosforilação
15.
ACS Appl Mater Interfaces ; 11(22): 19830-19840, 2019 Jun 05.
Artigo em Inglês | MEDLINE | ID: mdl-31088069

RESUMO

Tendon tissue engineering strategies that recreate the biophysical and biochemical native microenvironment have a greater potential to achieve regeneration. Here, we developed tendon biomimetic scaffolds using mechanically competent yarns of poly-ε-caprolactone, chitosan, and cellulose nanocrystals to recreate the inherent tendon hierarchy from a nano-to-macro scale. These were then coated with tropoelastin (TROPO) through polydopamine (PDA) linking, to mimic the native extracellular matrix (ECM) composition and elasticity. Both PDA and TROPO coatings decreased surface stiffness without masking the underlying substrate. We found that human adipose-derived stem cells (hASCs) seeded onto these TROPO biomimetic scaffolds more rapidly acquired their spindle-shape morphology and high aspect ratio characteristic of tenocytes. Immunocytochemistry shows that the PDA and TROPO-coated surfaces boosted differentiation of hASCs toward the tenogenic lineage, with sustained expression of the tendon-related markers scleraxis and tenomodulin up to 21 days of culture. Furthermore, these surfaces enabled the deposition of a tendon-like ECM, supported by the expression of collagens type I and III, tenascin, and decorin. Gene expression analysis revealed a downregulation of osteogenic and fibrosis markers in the presence of TROPO when compared with the control groups, suggesting proper ECM deposition. Remarkably, differentiated cells exposed to TROPO acquired an elastogenic profile due to the evident elastin synthesis and deposition, contributing to the formation of a more mimetic matrix in comparison with the PDA-coated and uncoated conditions. In summary, our biomimetic substrates combining biophysical and biological cues modulate stem cell behavior potentiating their long-term tenogenic commitment and the production of an elastin-rich ECM.


Assuntos
Adipócitos/citologia , Biomimética/métodos , Células-Tronco/citologia , Tendões/citologia , Tropoelastina/química , Diferenciação Celular/fisiologia , Proliferação de Células/fisiologia , Células Cultivadas , Matriz Extracelular , Humanos , Engenharia Tecidual
16.
J Dairy Sci ; 102(7): 6551-6554, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-31056330

RESUMO

Lameness and body condition are closely related. Recent studies have shown that cows with low body condition score (BCS) have a greater risk for developing lameness than cows with higher BCS. Among other reasons, this relationship might be related to the reduced thickness of the digital fat cushion in lean cows. The digital cushion is not a homogeneous structure but consists of different fat pads and connective tissue. We hypothesized that either high or low BCS will result in concordant adipocyte sizes in the fat pads of the digital cushion and subcutaneous tailhead fat irrespective of the localization of the latter. Right front claws were collected from 18 Holstein Friesian cows. Cows were selected according to their BCS: 9 cows with BCS <3 (low BCS) and 9 cows with BCS ≥3 (high BCS). After dissecting the horn capsule of the lateral claw, samples of the axial and abaxial fat pads were prepared for histomorphological examinations (adipocyte size measurement) and protein abundance of vascular endothelial growth factor A (VEGF-A) via Western blotting. In addition, fat samples were excised from the tailhead of all cows and used for the same purposes. Adipocyte size in tailhead fat was greater in high-BCS than in low-BCS cows. Similar differences between the BCS groups were apparent for adipocytes from the axial fat pad, although adipocytes in tailhead fat were larger than those in the digital cushion. In contrast to that in the axial fat pad and tailhead fat, adipocyte size in the abaxial fat pad was similar in cows from both BCS groups. A relationship between adipocyte size and VEGF-A protein was only confirmed for the axial fat pad, not for the other fat depots. When comparing BCS groups, differences in VEGF-A protein abundance between high-BCS and low-BCS cows were also limited to the axial fat pad, being absent in tailhead fat and the abaxial fat pad. Taken together, our results show that the fat pads from the digital cushion should not be considered uniform adipose tissue locations but rather discrete units reacting differently to fat mobilization.


Assuntos
Adipócitos/citologia , Tecido Adiposo/metabolismo , Fator A de Crescimento do Endotélio Vascular/metabolismo , Adipócitos/metabolismo , Tecido Adiposo/crescimento & desenvolvimento , Animais , Bovinos/crescimento & desenvolvimento , Bovinos/metabolismo , Tamanho Celular , Feminino , Gordura Subcutânea/crescimento & desenvolvimento , Gordura Subcutânea/metabolismo
17.
Food Funct ; 10(5): 2958-2969, 2019 May 22.
Artigo em Inglês | MEDLINE | ID: mdl-31073569

RESUMO

Cacao (Theobroma cacao) has a significant polyphenol content and has been reported to elicit anti-obesity effects. Previous studies have focused on the properties of cacao extract and procyanidins, while the potential mechanisms have not been fully elucidated. Here, we investigated the inhibitory effects of procyanidin metabolites on adipogenic cocktail-induced adipogenesis and lipogenesis in 3T3-L1 preadipocytes. It was observed that 5-(3',4'-dihydroxyphenyl)-γ-valerolactone (DHPV), a major procyanidin metabolite, exhibited the greatest inhibitory effects on adipogenesis and lipogenesis. DHPV dose-dependently reduced the expression levels of proteins involved in adipogenesis including peroxisome proliferator-activated receptor γ (PPAR γ) and CCAT/enhancer-binding protein α (C/EBP α), as well as lipogenesis-related factors such as fatty acid synthase and acetyl-CoA carboxylase. These inhibitory effects were primarily due to G1 phase arrest and the suppression of cell proliferation during mitotic clonal expansion, the early stage of adipogenesis. In an extensive kinase array, DHPV directly suppressed activation of the CDK2/cyclin O complex, and inhibited the phosphorylation of C/EBP ß, which is responsible for the induction of PPAR γ and C/EBP α. Taken together, these findings suggest that DHPV is a highly biologically active compound with potential anti-obesity effects and works by inhibiting the intracellular lipid content and cell differentiation.


Assuntos
Adipogenia/efeitos dos fármacos , Biflavonoides/metabolismo , Cacau/química , Catequina/metabolismo , Ciclo Celular/efeitos dos fármacos , Quinase 2 Dependente de Ciclina/metabolismo , Ciclinas/metabolismo , Lactonas/farmacologia , Proantocianidinas/metabolismo , Adipócitos/citologia , Adipócitos/efeitos dos fármacos , Adipócitos/metabolismo , Animais , Proteína alfa Estimuladora de Ligação a CCAAT/genética , Proteína alfa Estimuladora de Ligação a CCAAT/metabolismo , Quinase 2 Dependente de Ciclina/genética , Ciclinas/genética , Lactonas/metabolismo , Camundongos , Células NIH 3T3 , PPAR gama/genética , PPAR gama/metabolismo , Fosforilação
18.
Molecules ; 24(10)2019 May 23.
Artigo em Inglês | MEDLINE | ID: mdl-31126054

RESUMO

Dexamethasone is a glucocorticoid analog, which is reported to induce insulin resistance and to exacerbate diabetic symptoms. In this study, we investigated the association between mitochondrial dysfunction and the pathophysiology of dexamethasone-induced insulin resistance. An insulin resistance model in 3T3-L1 adipocyte was established by 48-h treatment of 1 µM dexamethasone, followed with the detection of mitochondrial function. Results showed that dexamethasone impaired insulin-induced glucose uptake and caused mitochondrial dysfunction. Abnormality in mitochondrial function was supported by decreased intracellular ATP and mitochondrial membrane potential (MMP), increased intracellular and mitochondrial reactive oxygen species (ROS) and mtDNA damage. Mitochondrial dynamic changes and biogenesis were suggested by decreased Drp1, increased Mfn2, and decreased PGC-1, NRF1, and TFam, respectively. The mitochondrial DNA (mtDNA) copy number exhibited no change while the mitochondrial mass increased. In agreement, studies in isolated mitochondria from mouse liver also showed dexamethasone-induced reduction of mitochondrial respiratory function, as suggested by decreased mitochondrial respiration controlling rate (RCR), lower MMP, declined ATP synthesis, opening of the mitochondrial permeability transition pore (mPTP), damage of mtDNA, and the accumulation of ROS. In summary, our study suggests that mitochondrial dysfunction occurs along with dexamethasone-induced insulin resistance in 3T3 L1 adipocytes and might be a potential mechanism of dexamethasone-induced insulin resistance.


Assuntos
Adipócitos/citologia , Dexametasona/efeitos adversos , Resistência à Insulina , Mitocôndrias Hepáticas/efeitos dos fármacos , Células 3T3-L1 , Trifosfato de Adenosina/metabolismo , Adipócitos/efeitos dos fármacos , Adipócitos/metabolismo , Animais , Células Cultivadas , Dano ao DNA , DNA Mitocondrial/efeitos dos fármacos , Modelos Animais de Doenças , Glucose/metabolismo , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Camundongos , Camundongos Endogâmicos ICR , Mitocôndrias Hepáticas/genética , Mitocôndrias Hepáticas/metabolismo , Dinâmica Mitocondrial/efeitos dos fármacos
19.
J Agric Food Chem ; 67(24): 6785-6791, 2019 Jun 19.
Artigo em Inglês | MEDLINE | ID: mdl-31141356

RESUMO

Obesity is a worldwide epidemic contributing to a higher risk of developing maladies such as type 2 diabetes, heart disease, and cancer. Shiya tea (leaves of Adinandra nitida), a traditional Chinese tea, is widely consumed due to its palatable flavor and various curative effects, such as reducing blood pressure and blood lipids, as well as anti-inflammation, etc. However, no relevant research on the antiobesity effects of Shiya tea has been reported. In particular, no health-benefiting compounds, other than flavonoids, in Shiya tea have been reported. Thus, 3T3-L1 preadipocytes have been used as a bioactivity-guided identification model to verify the inhibitory effects of Shiya tea on adipogenesis, as well as to identify antiadipogenic compounds. Four triterpenoid saponins (1-4), including one new compound (2α,3α-dihydroxyursolic acid 28- O-ß-d-glucopyranosyl ester, compound 1), and a flavonoid (5) have been identified using NMR (1D and 2D NMR) and liquid chromatography (LC)-MS techniques. Compound 1, the major antiadipogenic constituent with an IC50 value of 27.6 µg/mL, has been identified for the first time in Shiya tea. To understand the structure-activity relationship, three hydrolytic compounds (1s, 2s, and 5s) were obtained to provide an inhibitory effect on lipid accumulation during 3T3-L1 adipocyte differentiation. The inhibitory effect of the triterpenoid (1s) possessing no sugar group decreased significantly, while the flavonoid (5s) also without a sugar group showed increased activity. In addition, the hydroxyl group position may also play a role in inhibitory efficacy.


Assuntos
Adipócitos/efeitos dos fármacos , Medicamentos de Ervas Chinesas/química , Ericales/química , Folhas de Planta/química , Células 3T3-L1 , Adipócitos/citologia , Adipogenia/efeitos dos fármacos , Animais , Medicamentos de Ervas Chinesas/isolamento & purificação , Espectroscopia de Ressonância Magnética , Espectrometria de Massas , Camundongos
20.
Molecules ; 24(10)2019 May 17.
Artigo em Inglês | MEDLINE | ID: mdl-31108834

RESUMO

Clitoria ternatea (commonly known as blue pea) flower petal extract (CTE) is used as a natural colorant in a variety of foods and beverages. The objective of study was to determine the inhibitory effect of CTE on adipogenesis in 3T3-L1 preadipocytes. The phytochemical profiles of CTE were analyzed by liquid chromatography and tandem mass spectrometry (LC-MS/MS). Anti-adipogenesis effect of CTE was measured by using Oil Red O staining, intracellular triglyceride assay, quantitative real-time PCR and western blot analysis in 3T3-L1 adipocytes. Cell cycle studies were performed by flow cytometry. Lipolysis experiments were performed using a colorimetric assay kit. In early stages, CTE demonstrated anti-adipogenic effects through inhibition of proliferation and cell cycle retardation by suppressing expression of phospho-Akt and phospho-ERK1/2 signaling pathway. The results also showed that CTE inhibited the late stage of differentiation through diminishing expression of adipogenic transcription factors including PPARγ and C/EBPα. The inhibitory action was subsequently attenuated in downregulation of fatty acid synthase and acetyl-CoA carboxylase, causing the reduction of TG accumulation. In addition, CTE also enhanced catecholamine-induced lipolysis in adipocytes. These results suggest that CTE effectively attenuates adipogenesis by controlling cell cycle progression and downregulating adipogenic gene expression.


Assuntos
Adipócitos/citologia , Adipogenia/efeitos dos fármacos , Clitoria/química , Metabolismo dos Lipídeos/efeitos dos fármacos , Extratos Vegetais/farmacologia , Células 3T3-L1 , Adipócitos/efeitos dos fármacos , Adipócitos/metabolismo , Animais , Proteínas Estimuladoras de Ligação a CCAAT/genética , Proteínas Estimuladoras de Ligação a CCAAT/metabolismo , Proliferação de Células/efeitos dos fármacos , Regulação para Baixo , Flores/química , Regulação da Expressão Gênica/efeitos dos fármacos , Lipólise , Camundongos , PPAR gama/genética , PPAR gama/metabolismo , Extratos Vegetais/isolamento & purificação
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