Cloning and expression profile of ZFP36L1 gene in goat / 生物工程学报

The purpose of this study was to clone and characterize the ZFP36L1 (zinc finger protein 36-like 1) gene, clarify its expression characteristics, and elucidate its expression patterns in different tissues of goats. Samples of 15 tissues from Jianzhou big-eared goats, including heart, liver, spleen, lung and kidney were collected. Goat ZFP36L1 gene was amplified by reverse transcription-polymerase chain reaction (RT-PCR), then the gene and protein sequence were analyzed by online tools. Quantitative real-time polymerase chain reaction (qPCR) was used to detect the expression level of ZFP36L1 in intramuscular preadipocytes in different tissues and adipocytes of goat at different differentiation stages. The results showed that the length of ZFR36L1 gene was 1 224 bp, and the coding sequence (CDS) region was 1 017 bp, encoding 338 amino acids, which was a non-secretory unstable protein mainly located in nucleus and cytoplasm. Tissue expression profile showed that ZFP36L1 gene was expressed in all selected tissues. In visceral tissues, the small intestine showed the highest expression level (P < 0.01). In muscle tissue, the highest expression level was presented in longissimus dorsi muscle (P < 0.01), whereas the expression level in subcutaneous adipose tissue was significantly higher than that in other tissues (P < 0.01). The results of induced differentiation showed that the expression of this gene was up-regulated during adipogenic differentiation of intramuscular precursor adipocytes (P < 0.01). These data may help to clarify the biological function of the ZFP36L1 gene in goat.

Equisetin is an anti-obesity candidate through targeting 11β-HSD1

Obesity is increasingly prevalent globally, searching for therapeutic agents acting on adipose tissue is of great importance. Equisetin (EQST), a meroterpenoid isolated from a marine sponge-derived fungus, has been reported to display antibacterial and antiviral activities. Here, we revealed that EQST displayed anti-obesity effects acting on adipose tissue through inhibiting adipogenesis in vitro and attenuating HFD-induced obesity in mice, doing so without affecting food intake, blood pressure or heart rate. We demonstrated that EQST inhibited the enzyme activity of 11β-hydroxysteroid dehydrogenase type 1 (11β-HSD1), a therapeutic target of obesity in adipose tissue. Anti-obesity properties of EQST were all offset by applying excessive 11β-HSD1's substrates and 11β-HSD1 inhibition through knockdown in vitro or 11β-HSD1 knockout in vivo. In the 11β-HSD1 bypass model constructed by adding excess 11β-HSD1 products, EQST's anti-obesity effects disappeared. Furthermore, EQST directly bond to 11β-HSD1 protein and presented remarkable better intensity on 11β-HSD1 inhibition and better efficacy on anti-obesity than known 11β-HSD1 inhibitor. Therefore, EQST can be developed into anti-obesity candidate compound, and this study may provide more clues for developing higher effective 11β-HSD1 inhibitors.

Cambios en el perfil proteómico de preadipocitos con diferentes condiciones metabólicas / Changes in the proteomic pro/ile of preadipocytes with different metabolic status

Resumen Introducción y objetivos: El tejido adiposo subcutáneo se considera un depósito con un papel protector desde un punto de vista metabólico. El exceso de tejido adiposo desencadena en obesidad, la cual, está acompañada típicamente por resistencia a insulina, dislipidemia, e hipertensión arterial. No obstante, se conoce que existe un subgrupo de obesos que parecen estar protegidos de dichas complicaciones. Estos individuos son definidos como obesos sanos metabólicamente. A pesar de los avances en el conocimiento de las alteraciones que suceden en el tejido adiposo en obesidad, aún se desconocen los mecanismos que subyacen en el desarrollo de resistencia a insulina. Por lo tanto, en este trabajo, se estudió la asociación entre obesidad y desarrollo de enfermedad metabólica identificando factores y procesos que determinan la transición desde el fenotipo obeso sano y no sano, empleando preadipocitos provenientes de tejido adiposo subcutáneo. Metodología: Se emplearon datos de un estudio de proteómica comparada de preadipocitos de tejido subcutáneo obtenidos de pacientes obesos normoglucémicos no resistentes a insulina y de pacientes obesos con diabetes mellitus de tipo 2. El estudio proteómico, se llevó a cabo utilizando la técnica de iTRAQ combinada con LC-MSMS. Resultados y conclusiones: Las diferencias entre preadipocitos de tejido adiposo subcutáneo en sujetos normoglucémicos y con diabetes, afectan sobre todo a proteínas citosólicas y, en particular, a proteínas relacionadas con procesos metabólicos mientras que, las membranales no cambian entre fenotipos obesos. En el estudio se identificaron importantes diferencias en el perfil proteómico de los preadipocitos de tejido adiposo subcutáneo en obesidad, tanto en sujetos normoglucémicos como diabéticos, apoyando la importancia de estas células en el mantenimiento de la identidad del depósito graso. También se encontró que, la transición desde el fenotipo obeso sano hacia el no sano conlleva un mayor desarrollo de estrés oxidativo e inflamación en las células precursoras adipocitarias.

Abstract Introduction and objectives: The subcutaneous adipose tissue is considered as a depot with a protective role from a metabolic point of view. An excess of adipose tissue is triggered in obesity, which is accompanied by insulin resistance, dyslipidemia and arterial hypertension. However, it is known that, there is a subgroup of obese people who seem to be protected from obese complications. These individuals are defined as metabolically healthy obese. Despite the advances in the knowledge of the alterations that occur in adipose tissue during obesity, the mechanisms underlying the development of insulin resistance are still unknown. Therefore, in this work, we studied the association between obesity and the development of metabolic disease, we identified factors and processes that determined the transition of healthy and unhealthy obesity phenotype, using preadipocytes from subcutaneous adipose tissue. Methods: Data obtained from a comparative proteomics study of subcutaneous adipose tissue preadipocytes from normoglycemic obese patients-not resistant to insulin and from obese patients with type 2 diabetes mellitus were used. The proteomic study was carried out using the iTRAQ combined with LC -MSMS. Results and conclusions: The differences between pre-adipocytes of subcutaneous adipose tissue in normoglycemic subjects and with diabetes affect mainly cytosolic proteins and, in particular, proteins related to metabolic processes while, membrane proteins do not change between obese phenotypes. In this study, we identified significant differences in the proteomic profile of preadipocytes from subcutaneous adipose tissue in obesity in both, normoglycemic and diabetic subjects, supporting the importance of these cells in the maintenance of the fat depot identity. We also found that, the transition from unhealthy to healthy phenotype in obesity, leads to further development of oxidative stress and inflammation in adipocyte precursor cells.

Effects of Lonicera caerulea extract on adipocyte differentiation and adipogenesis in 3T3-L1 cells and mouse adipose-derived stem cells (MADSCs) / 한국영양학회지

PURPOSE: Obesity is a major health problem of global significance because it is clearly associated with an increased risk of health problems, such as nonalcoholic fatty liver disease (NAFLD), diabetes, cardiovascular diseases, and cancer. Lonicera caerulea (LC) originates from high mountains or wet areas and has been used as a traditional medicine in northern Russia, China, and Japan. LC contains a range of bioactive constituents, such as vitamins, minerals, and polyphenols. This study examined the anti-obesity effects of LC during differentiation in preadipocytes. METHODS: The cell viability assay was performed after the differentiation of 3T3-L1 cells for 7 days. Oil Red O staining was used to visualize the changes in lipid droplets in 3T3-L1 cells and mouse adipose-derived stem cells (MADSCs). The mRNA expression of obesity-related genes was determined by quantitative real-time PCR. RESULTS: According to the results of Oil Red O staining, the lipid levels and size of lipid droplets in the adipocytes were reduced and the LC extract (LCE, 0.25–1 mg/mL) markedly inhibited adipogenesis in a dose-dependent manner. The treatment of LCE also decreased the mRNA expression of peroxisome proliferator-activated receptor γ (PPARγ), CCAAT/enhancer binding protein-α (C/EBPα), and sterol regulatory element binding protein 1 (SREBP1) in 3T3-L1 cells. Western blot analysis showed that the PPARγ, C/EBPα, and SREBP1 protein levels in both 3T3-L1 and MADSC were reduced in a dose-dependent manner. CONCLUSION: These results suggest that LCE can inhibit adipogenic differentiation through the regulation of adipogenesis-related markers.

miR-202 promotes the differentiation of 3T3-L1 preadipocyte via inhibiting PGC1β expression / 基础医学与临床

Objective To explore the effect and molecular mechanism of miR-202 on the differentiation of 3T3-L1 preadipocyte .Methods Through lentivirus infected with 3T3-L1 preadipocytes , we set up the AMO-miR-202 group and the random control group , then, these cells were induced to differentiate , nine days later, differentiation was assessed by Oil Red O staining and we examined the mRNA expression of PPARγ2 and aP2 by RT-PCR method. We examined the mRNA expression of PPARγ2,aP2 and PGC1βby Western blot method .Results After packa-ging lentivirus with AMO-miR-202 and random sequence control miRNA through cell line 293T, 80%-90%cells with fluorescence were found under fluorescence microscope; After these two lentivirus respectively infected with 3T3-L1 preadipocytes, About 70%-80%cells with fluorescence were found under fluorescence microscope .Oil Red O staining test showed that these cells with Oil Red O stained bright red fat droplets of AMO-miR-202 group and PPARγ2 and aP2 mRNA expression in the AMO-miR-202 group significantly lower than control groups (P<0.05). Western blot assay showed that the protein expression of PGC 1βin the AMO-miRNA-202 group was significantly increased(P<0.05), but the expression of aP 2 and PPARγ2 was significantly decreased (P<0.01).However, the random control group and the adipocyte group had no significant effect on the above indexes .Conclusions miR-202 can promote the differentiation of 3T3-L1 preadipocyte by inhibiting the protein expression of PGC 1βand im-proving the protein expression of PPARγ2 and aP2.

The Role of Proliferation and Differentiation of Preadipocyte on High Fat Diet-induced Obesity and Prevention of Obesity through Exercises in Rats / 中国运动医学杂志

Objective To explore the influences of proliferation and differentiation of preadipocytes on high fat diet-induced obesity and obesity prevention through exercises in rats.Methods Forty male Sprague-Dawley rats were divided into a normal diet control group (C,n=8),a normal diet with exercises group (E,n=8),a high fat diet control group (H,n=14) and a high fat diet with exercises group (HE,n=10).Group C and group H kept sedentary,while group E and group HE underwent treadmill exercises at about 75％VO2max level.After 12 weeks of intervention,the body weight,epididymal,perirenal and retroperitoneal fat mass were recorded,and the total fat mass was determinated.Stereology method was used to calculate the number and average diameter of fat cells.Western Blotting was conducted to measure the expression of PPARγ and C/EBPα protein in adipose tissues.Results (1) The body weight and total fat mass of group H were significantly higher than those of group C (P＜0.01).Perirenal,retroperitoneal fat and total fat mass of group E and HE were significantly lower than those of group C and H respectively (P＜0.01).(2) The total fat cell number of group H was significantly higher than that of group C.The average diameter of fat cells in perirenal and retroperitoneal fat pads of group E were significantly lower than that of group C (P＜0.05,P＜0.01),while that of group HE in epididymal,perirenal and retroperitoneal fat pads was significantly lower than that of group H (P＜0.01).(3) Compared with group C,the expression of PPARγ protein of group E and H increased significantly in epididymal,perirenal and retroperitoneal fat pads (P＜0.01).The expression of C/EBPα protein of group H was significantly higher than that of group C in epididymal and perirenal fat pads (P＜0.01),the expression of C/EBPα protein of group E was significantly higher than that of group C in perirenal and retroperitoneal fat pads (P＜0.01),while the expression of C/EBPα protein of group HE was significantly lower than that of group H in perirenal fat pads(P＜0.01).Conclusion The proliferation and differentiation of preadipocyte was enhanced in the development of high fat diet induced obesity and exercises to prevent obesity,but its role and mechanisms were different.The high fat diet increases the number of fat cells which is a compensatory mechanism for the body to adapt to fat accumulation,while exercises might promote cell renewal and decrease the average size of fat cells which is an adaptive mechanism to improve fat storage.

Serum Preadipocyte Factor 1 Levels Are Not Associated with Bone Mineral Density among Healthy Postmenopausal Korean Women

BACKGROUND: Multipotent mesenchymal stem cells can differentiate into adipocytes or osteoblasts through closely regulated lineage-control processes. However, adipocyte precursor cells release preadipocyte factor 1 (Pref-1), which inhibits the differentiation of mesenchymal stem cells into mature adipocytes and osteoblasts. Previous studies have also reported an inverse association between Pref-1 levels and bone mineral density (BMD) among patients with anorexia nervosa. METHODS: In this retrospective study, we examined the correlations between Pref-1 levels and BMD among 124 healthy postmenopausal women (>50 years old). The patients had provided information regarding their clinical characteristics, and underwent blood testing and serum Pref-1 testing. RESULTS: The subjects' mean age was 59.9±7.1 years and the median time since menopause onset was 9.1 years. A history of osteoporotic fracture was identified in 23 subjects (19%). Serum Pref-1 levels were not significantly correlated with BMD values at the lumbar spine (R²=0.038, P=0.109), femur neck (R²=0.017, P=0.869), and total hip (R²=0.041, P=0.09), and multivariate analyses with adjustment for age and body mass index also did not detect any significant correlations. Subgroup analyses according to a history of fracture also did not detect significant associations between Pref-1 levels and BMD values. CONCLUSION: In our study population, it does not appear that serum Pref-1 levels are significantly associated with BMD values and osteoporosis.

El adipocito en el laboratorio: historia, perspectivas y reporte de experiencia / The adipocyte in the laboratory: history, perspectives and report of experience

Introducción: el tejido adiposo humano está compuesto por diferentes tipos celulares, y ha sido objeto de múltiples estudios en los últimos años debido a su acción en diversas funciones. Métodos: se realizó una búsqueda bilbiográfica en PubMed, Scielo, Science Direct and Google Scholars y se reporta la experiencia de los autores. Resultados: los primeros modelos de estudio fueron con tejido de roedores, y permitieron la comprensión del metabolismo de carbohidratos y ácidos grasos y facilitaron el estudio de la biología del adipocito, junto a estudios histológicos y el aislamiento de adipocitos maduros. Posteriormente se realizaron estudios con células precursoras (preadipocitos) que propiciaron el aislamiento de la línea celular 3T3-1L murina. En Latinoamérica, se han realizado diversos estudios con líneas celulares y con células madre mesenquimales precursoras de adipocitos para estudiar el efecto de hormonas y otras sustancias y para genotipificación. En Colombia se han realizado estudios con adipocitos 3T3-L1 para determinar los efectos de medicamentos y sustancias en estas células. En el laboratorio de Fisiología Celular de la Universidad Tecnológica de Pereira el proceso de obtención de muestras ha evidenciado dificultades por tratarse de tejido humano, pero el protocolo de aislamiento y cultivo pudo ser estandarizado a lo largo de seis años de experimentación; se aislaron preadipocitos y adipocitos maduros que permitieron estudiar los efectos de hormonas, realizar caracterización electrofisiológica y estudiar la fisiología del calcio. Conclusiones: este es un campo de investigación muy relevante debido a la implicación de este tipo celular en funciones metabólicas sistémicas y su relación con patologías de alta prevalencia como la obesidad y el síndrome metabólico.

Introduction: The human adipose tissue is composed of different cell types. It has been subject of several studies in the past years due to its role on diverse functions. Methods: A search in the PubMed, Scielo, Science direct and Google Scholars databases was performed and the authors experience was reported. Results: The first model used was rodent fat, which allowed a better comprehension on carbohydrate and fatty acid metabolism and enhanced research in adipocyte cell biology in addition with histological studies and mature adipocyte isolation. Afterwards, learning about precursor cell (pre-adipocytes) promoted the isolation of murine 3T3-L1 cell line. In Latin America research has been conducted using cell lines and adipocyte precursor mesenchymal stem cells to describe effects of hormones and perform DNA sequencing. In Colombia, studies in 3T3-L1 cell line aimed to stablish the effects of different compounds on these cells. In the Cell Physiology Laboratory of the Universidad Tecnológica de Pereira, sample collection process has shown difficulties because the source was human tissue; nevertheless isolation and cell culture protocols were standardized throughout the last six years of experimentation. Pre-adipocytes and mature adipocytes were isolated to study the effects of hormones, perform electrophysiological characterization and study calcium physiology. Conclusions: This is a relevant research field since these cells have important systemic metabolic functions and they have a clear relationship with high-prevalence pathologies such as obesity and the metabolic syndrome.

Effects of 3'-hydroxygenistein on proliferation and differentiation of 3T3-L1 preadipocytes and the underlying mechanisms / 第二军医大学学报

Objective To explore the effects of 3'-hydroxygenistein on the proliferation and differentiation of 3T3-L1 preadipocytes and to elucidate the related mechanism. Methods 3T3-L1 preadipocytes were cultured and treated with different dosages of 3'-hydroxygenistein. Cell proliferation was analyzed by counting cell numbers and MTT assay; DNA synthesis of 3T3-L1 was investigated by bromodeoxyuridine (BrdU) incorporation assay; the degree of preadipocytes differentiation was evaluated by Oil red O staffing; and the expressions of peroxisome proliferation activated receptor γ (PPARγ) and CAAT/ enhancer binding protein (C/EBPα) were detected at mRNA and protein level by real-time PCR and Western blolting analysis, respectively. Results Pretreatment with 3'-hydroxygenistrm (10-50 μmol/L) for 24, 48 and 72 h markedly promoted 3T3-L1 preadipocyte proliferation and differentiation in a dose-effect manner (P<0. 05, P<0. 01). It also significantly facilitated the DNA synthesis in 3T3-L1 preadipocytes in a dose-dependent manner (P< 0. 05). Similar to rosiglitazone (ROZ), 3'- hydroxygenistein (at 10 or 50 μmol/L) also resulted in more lipid droplets in 3T3-L1 preadipocytes (P<0. 05, P<0. 01). Furthermore, 3'-hydroxygenistein greatiy increased the mRNA and protein expression of PPARγ and C/EBPα in 3T3-L1 preadipocytes (P<0. 01). Conclusion 3'-hydroxygenistein can promote the proliferation and DNA synthesis of 3T3-L1 preadipocytes; it can also enhance the accumulation of lipid drops and increase the terminal differentiation of preadipocyts, which might be associated with its ablity to increase the expression of PPARγ and C/EBPα.

Role of miR-24 in the regulation of 3T3-L1 adipicyte differentiation and the expression of fatty acid binding protein / 中国药理学通报

Aim To explore the effect of microRNAs on the differentiation of 3T3-L1 adipocytes and the expression of adipo-related gene-fatty acid binding protein during the adipocyte differentiation.Methods adipo-related microRNAs during 3T3-L1 adipocyte differentiation were screened and identified by micorRNA microarray.Constructed high-expression plasmids of the adipo-related microRNAs,were transfected into the 3T3-L1 preadipocytes by lipofectamine.While the effect of adipo-related microRNAs on the course of 3T3-L1 adipocyte differentiation was observed,the protein and mRNA expression level of fatty acid binding protein(FABP4)were analyzed by Western blot and RT-PCR during 3T3-L1 adipocyte differentiation.Results The expression profiles of microRNAs have significant changed during 3T3-L1 adipocyte differentiation,in which 35 microRNAs among them down-relation,the most lowly expression is miR-24;17 microRNAs among them up-relation,the most highly expression is miR-21.MiR-24 significantly inhibited adipocyte differentiation and maturity,while miR-21 have no significant effect.MiR-24 significantly inhibited the expression of FABP4,but had no effect on the level of its mRNA;miR-21 had no effect on the expression of protein and mRNA of FABP4.Conclusion There exist adipogenic-related microRNAs during 3T3-L1 adipocyte differentiation; miR-24 play an important role in the regulation of 3T3-L1 preadipocyte differentiation into adipocyte and the(FABP4)protein expression.

Obestatin inhibits proliferation and differentiation of 3T3-L1 preadipocytes / 第二军医大学学报

Objective: To investigate the inhibitory effects of obestatin on proliferation and differentiation of 3T3-L1 preadipocytes. Methods: Obestatin (10-8 mmol/L, 10-9 mmul/L, 10-10 mmol/L, 10-11 mmol/L, and 10-12 mmol/L) and ghrelin (10-10 mmol/L) were used to treat 3T3-L1 preadipocytes. Cell proliferation was assessed by MTT assay and the results were compared with that of blank control group. The differentiation of 3T3-L1 preadipocytes (from day 1 to day 10) was interfered with obestatin or ghrelin (both at 10-10 mmol/L). Intracellular fat accumulation in differentiated adipocytes was determined by oil red O staining and the expression of PPAR-γ2 mRNA was detected by RT-PCR; the results were compared with that of control group (induced with routine inducer). Results: Compared with the blank control group, obestatin-treated groups (various concentrations of obestatin) had significantly less cells(P<0.05). Oil red O staining revealed that, compared with control group, the formation of lipid droplets was significantly decreased after 10 days' of treatment with 10-10 mmol/L obestatin (P<0.05). The expression of PPARγ2 gene increased with the progress of 3T3-L1 preadipocytes differentiation. PPAR-γ2 mRNA level in mature adipocytes of obestatin group was significantly lower than that in the cells of control group. The effect of ghrelin was contrary to that of obestatin. Conclusion: Obestatin can inhibit the proliferation and differentiation of 3T3-L1 preadipocyes.

Effect of rh-leptin on the differentiation of orbital preadipocytes in thyroid associated ophthalmopathy / 眼科研究

Background Thyroid-associated ophthalmopathy (TAO) is characterized by increased amount of orbital fat tissue.Leptin is a specific adipocytokine,and it may play an important role in the pathogenesis of TAO.Objective The present study investigates the effect of rh-leptin on the differentiation of orbital preadipocytes in subjects suffering from thyroid-associated ophthalmopathy (TAO).Methods The orbital fatty tissue was obtained from 7 patients with TAO during orbital decompression surgery.Orbital preadipocytes were cultured using the explant method and differentiated cells were identified with oil red staining.The subcultured preadipocytes were incubated with different concentrations of rh-leptin,and no rh-leptin was added in the control group.The average optical density of cytoplasm/nucleolus in differentiated adipocytes was measured by oil-red staining to evaluate the influence of different concentrations of rh-leptin on lipid formation of orbital adipocytes.Results Cultured preadipocytes in vitro were identified to be of mesenchymal origin by immunohistochemical staining.The average optical density of cytoplasm/nucleolus was 1.07±0.22,0.80±0.17,0.56±0.11,0.25±0.10,and 0.17±0.08,respectively in the control group,10μg/L of leptin group,50μg/L of leptin group,100μg/L of leptin group and 500μg/L of leptin group,showing a significant difference among the five groups (F=20.64,P=0.00).Compared to the control group,the lipid formation of orbital adipocytes gradually declined in various concentrations of rh-leptin (P＜0.05,P＜0.01).Conclusion Rh-leptin may suppress the differentiation and lipid formation of TAO orbital preadipocytes in vitro in a dose-dependent manner.Leptin may be a feedback modulator in TAO pathogenesis.

Effects of leptin on proliferation and differentiation of human preadipocyte / 中华医学美学美容杂志

Objective To observe the effects of leptin on the proliferation and differentiation of human preadipocyte in vitro, and to explore the possible mechanism of the generation of obesity regulated by leptin. Methods The human preadipocytes were isolated from human subcutaneous adipose tissue of abdomen and cultured in vitro. The effects of leptin (0-1 000 ng/ml) on the proliferation, lipid accumulation and the mRNA expression of PPAR-γ2 and C/EBP-α, which are the differentiation and transcription factor of human preadipocyte, were analyzed by the methods of MTT, cell counting, extracting stained intracytoplasmic lipid with oil red O and RT-PCR. Results Leptin (1 000 ng/ml) could stimulate the proliferation, lipid accumulation and the mRNA expression of PPAR-γ2 and C/EBP-α (P<0. 05). There were not obvious effects on the proliferation and lipid accumulation in the groups of lower (10 ng/ml) and common (100 ng/ml) concentration (P>0. 05). Conclusion Leptin in higher concentration can stimulate the proliferation and differentiation of preadipocye in vitro, which indicates that leptin may regulate the generation of obesity through acting on the proliferation and differentiation of preadipocyte at the pathologic state of leptin resistance and high leptin concentration in serum.

Characterization of gene expression and adipokine secretion during differentiation of primary cultured human omental preadipocytes / 中华内分泌代谢杂志

Stromal preadipocytes from human omental adipose tissues were isolated with enzyme-digesting method,and cultured for inducing differentiation.Preadipocyte factor-1 was abundant during preadipoeyte stage,whereas lipoprotein lipase and PPARγ2 mRNA were rapidly up-regulated at early stage of differentiation and CCAAT/enhancer binding protein expression was increased rather late.In addition,resistin expression and secretion was primarily detectable in preadipocytes,leptin was gradually increased during adipogenesis,whereas adiponectin appeared specifically in mature adipocytes.Insulin-like growth factor-Ⅰ secretion was remarkably detected in both preadipocytes and mature adipocytes.

Effects of Different Scaffolds and Cultures on Growth of Preadipocytes / 中华医学美学美容杂志

Objective To culture preadipocytes in vitro and to study the cell compatibility of PLGA scaffolds,collagen scarfolds and hyaluronic acid-based scaffolds and tO choose the optimal seeding method.Methods The preadipocytes from human abdominal adipose tissue were isolated and cultured in enzyme-digesting method.The generation of human preadipocytes was planted on PLGA scaffolds,collagen scaffolds and hyaluronic acid-based scaffolds.and the cell compatibility was observed by MTT method.The seeding efficiency of human preadipocytes on scaffolds.human preadipocytes were seeded to hyaluronic acid-based scaffolds by static culture and stirred culture.Results Compared compatibility of preadipocyte with three different scaffolds,there was great difference between hyaluronic acid-based scaffolds and PLGA scaffolds.Difference also existed between hyaluronic acid-based scaffolds and collagen scaffolds that were different from PLGA scaffolds.Among them,hyaluronic acid-based scaffolds was the best.Conclusion Hyaluronic acid iS a better scaffolds material for adipose tissue engineering compared with PLGA and collagen.The seeding efficiency of stirred culture is higher than static culture,which is an optimal method for cell seeding tO 3-D scaffolds.

Effects of Ghrelin on the Proliferation and Differentiation of 3T3-L1 Preadipocytes / 华中科技大学学报（医学）（英德文版）

The effects of ghrelin on the proliferation and differentiation of 3T3-L1 preadipocytes and the possible mechanisms were investigated in this study.3T3-L1 preadipocytes were cultured in vitro and treated with different concentrations of ghrelin.Proliferation of 3T3-L1 preadipocytes was evaluated by MTT method and mRNA levels of c-myc and thymidine kinase were detected by RT-PCR.Morphological changes of 3T3-L1 preadipocytes were observed and cell differentiation was measured by oil red O staining.The mRNA levels of peroxisome proliferator-activated receptor γ (PPARγ) and CAAT/enhancer binding protein (C/EBPα) in the cells at different differentiation stages were detected by RT-PCR.The results showed that ghrelin at concentrations of 10-7 to 10-15 mol/L could significantly promote preadipocyte proliferation (P<0.05),with the most pronounced effect observed at 1011mol/L (P<0.01).Treatment of 3T3-L1 preadipocytes with ghrelin significantly in-creased the mRNA levels of c-myc and thymidine kinase (P<0.01).Morphological findings demonstrated that the great amount of lipid droplets appeared in the 3T3-L1 preadipocytes treated with ghrelin.Ghrelin could morphologically induce the differentiation of 3T3-L1 preadipocytes into mature adipocytes.Ghrelin significantly increased the mRNA levels of PPART and C/EBPα during the differentiation,when compared with control group (P<0.05).The mRNA levels of PPARγ and C/EBPα were obviously up-regulated with the differentiation of preadipocytes after the treatment of ghrelin.There were significant difference in the mRNA levels of PPARγ and C/EBPα on day 2 and day 8 of the differentiation of 3T3-L1 preadipocytes (P<0.01).In conclusion,ghrelin could promote the proliferation and differentiation of 3T3-L1 preadipocytes by increasing the mRNA levels of PPARγ and C/EBPα and therefore enhance the sensitivity of adipocytes against insulin.

Leptin and lipogenesis / 国际外科学杂志

Leptin,the product of the obese gene,is a protein produced and secreted primarily by adipose tissue.Leptin initiates several biological effects by binding to its receptors.Many studies have shown that leptin can reduce fat accumulation,but its mechanism is still unclear.This article reviewed the effects of leptin on lipogenesis,and especially on proliferation and differentiation of preadipocytes.

Primary culture of human preadipocyte in a serum-free medium / 基础医学与临床

Objective To establish a method of primary culture of human preadipocyte in a serum-free medium. Methods Collangenase digestion was used to dispart preadipocyte.The cells were identified by microscopy stained by oil red O and determination of the activity of Glycerol-3-phosphate dehydronase(G-3-PDH). Results The primary cultured human preadipocyte proliferated in the aserum-free medium successfully.In the differential serum-free medium the cells turned to be round on the 4 th day.The adipose drops began to cumulate in the cells,and to the most quantity until the 21 st day.Conclusion The human preadipocyte can be primarily cultured and induced to differentiate in serum-free medium,which is the base for researching the effects of hormones and factors to the proliferation and differentiation of preadipocyte.

The Interaction of Adipose Tissue with Immune System and Related Inflammatory Molecules

BACKGROUND: Adipose tissues were initially introduced as energy storages, but recently they have become famous as an endocrine organ which produces and secretes various kinds of molecules to make physiologic and metabolic changes in human body. It has been studied that these molecules are secreted in abundance as the adipose tissue becomes bigger along with obesity. Furthermore, it has been found that they are mediating systemic inflammation and generation of metabolic diseases such as type 2 diabetes and atherosclerosis. On the basis of these, we studied previous papers which have been researched about the interaction between preadipocytes and macrophages, adipose tissues and lymph nodes, and adipose tissue secreting molecules. RESULTS: Firstly, preadipocytes and macrophages are expressing similar transcriptomes and proteins, and preadipocytes can be converted to mature macrophages which have phagocytic activity. Moreover, the monocytes, which initially located in the bone marrow, are filtrated to the adipose tissue by monocyte chemotatic protein-1 and are matured to macrophages by colony stimulating factor-1. Secondly, adipose tissues and their associated lymph nodes are interacting each other in terms of energy efficiency. Lymph nodes promote lipolysis in adipose tissues, and polyunsaturated fatty acids in adipocytes become energy sources for dendritic cells. Lastly, adipose tissues produce and secrete proinflammatory molecules such as leptin, adiponectin, TNF-alpha, IL-6, and acute phase proteins, which induce the inflammation and potentially generate metabolic diseases. CONCLUSION: According to these, we can link adipose tissues to inflammation, but we need to affirm the actual levels and roles of adipose tissue-derived proinflammatory molecules in human body.

Effects of Arachidonic Acid on Growth and Differentiation of Rat Preadipocytes / 中国生物化学与分子生物学报

To study effects of arachidonic acid (AA) on the growth and differentiation of rat adipocytes, a cells culture system of rat primary preadipocytes was established. The cells treated by different concentration of AA supplemented based on DMEM medium containing 10% fetal bovine serum. Cell proliferation was measured by trypan blue exclusion and methyl thiazolyl tetrazolium (MTT) assay method. Hoechst33342 fluorescence staining observed AA induced morphological changes. Oil Red O staining extraction assay assess the degree of adipogenesis and differentiation, and cyclooxygenases-2(COX-2) mRNA were detected by reverse transcriptase-polymerase chain reaction (RT-PCR). The results showed that rat preadipocytes treated with 120 μmol/L AA for 24-72 hours remarkably promoted the cells proliferation compared with control, 160 μmol/L AA treated for 48 hours could induce apoptosis of preadipocytes. 40, 80 μmol/L AA decreased the fat content in cells at 72 hours, and 40 μmol/L AA significantly up-regulated the expression of COX-2 mRNA at 24 hours. This results indicate that AA regulate adipocytes proliferation and differentiation depended on treatment time and concentration. 40-80 μmol/L AA maybe useful to control body fat, which may be associated with the increase of COX-2 mRNA.