• Title/Summary/Keyword: mitotic clonal expansion

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Inhibitory Effects and Molecular Mechanism of Adipocyte Differentiation by Rosae laevigata Fructus Ethanol Extracs (금앵자 에탄올 추출물에 의한 3T3-L1 지방세포의 분화억제 효과와 그 메커니즘 규명)

  • Jeong, Hyun Young;Jeong, In Kyo;Nam, So Yeon;Yun, Hee Jung;Kim, Byung Woo;Kwon, Hyun Ju
    • Microbiology and Biotechnology Letters
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    • v.44 no.1
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    • pp.89-97
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    • 2016
  • Obesity is caused by excess accumulation of body fat and contributes to various pathological disorders such as diabetes, hypertension, cardiovascular disease, and cancer. In this study, we investigated the effect of a 30% ethanol extract of Fructus Rosae laevigata (RLE) on adipogenesis in 3T3-L1 adipocytes, measured by triglyceride accumulation and expression of adipogenesis-related transcription factors during differentiation of pre-adipocytes into adipocytes. RLE decreased the intracellular triglyceride contents (assessed by Oil Red-O staining) in a dose-dependent manner. It also downregulated the expression of adipogenic transcription factors and inhibited cell proliferation during the mitotic clonal expansion phase of adipocyte differentiation by inducing G1 phase arrest. We investigated the alterations in the levels of G1 phase arrest-related proteins. The expression of p21 protein significantly increased, while the levels of Cyclin E, Cdk2, and phospho-Rb decreased in a dose-dependent manner in 3T3-L1 cells treated with RLE. These results suggest that RLE inhibits the differentiation of 3T3-L1 adipocytes by suppressing the expression of adipogenic transcription factors and inducing G1 phase arrest in the early stages of adipocyte differentiation.

Inhibition of Adipocyte Differentiation by MeOH Extract from Carduus crispus through ERK and p38 MAPK Pathways

  • Lee, Eun-Jeong;Joo, Eun-Ji;Hong, Yoo-Na;Kim, Yeong-Shik
    • Natural Product Sciences
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    • v.17 no.4
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    • pp.273-278
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    • 2011
  • In this study, the effects of a methanol (MeOH) extract of Carduus crispus L. (Asteraceae) on adipogenesis was investigated in 3T3-L1 cells. To differentiate preadipocytes to adipocytes, confluent 3T3-L1 preadipocytes were treated with a hormone mixture, which included isobutylmethylxanthine, dexamethasone, and insulin (MDI). The methanol extract of C. crispus significantly decreased fat accumulation by inhibiting adipogenic signal transcriptional factors in MDI-induced 3T3-L1 cells in a dose-dependent manner. In MTT assays and on PI-staining, methanol extract of C. crispus inhibited the proliferation of 3T3-L1 cells during mitotic clonal expansion (MCE). The anti-adipogenic effect of the Carduus extract seemed to be associated with the upregulation of extracellular signal-regulated kinase (ERK) and p38 mitogen-activated protein kinase (MAPK) pathways within the first 2 days after MDI treatment. These results suggest that methanol extract of C. crispus might be beneficial for the treatment of obesity.

Lignosulfonic acid promotes hypertrophy in 3T3-L1 cells without increasing lipid content and increases their 2-deoxyglucose uptake

  • Hasegawa, Yasushi;Nakagawa, Erina;Kadota, Yukiya;Kawaminami, Satoshi
    • Asian-Australasian Journal of Animal Sciences
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    • v.30 no.1
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    • pp.111-118
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    • 2017
  • Objective: Adipose tissue plays a key role in the development of obesity and diabetes. We previously reported that lignosulfonic acid suppresses the rise in blood glucose levels through the inhibition of ${\alpha}$-glucosidase activity and intestinal glucose absorption. The purpose of this study is to examine further biological activities of lignosulfonic acid. Methods: In this study, we examined the effect of lignosulfonic acid on differentiation of 3T3-L1 cells. Results: While lignosulfonic acid inhibited proliferation (mitotic clonal expansion) after induction of differentiation, lignosulfonic acid significantly increased the size of accumulated lipid droplets in the cells. Semi-quantitative reverse transcription polymerase chain reaction analysis showed that lignosulfonic acid increased the expression of the adipogenic transcription factor, peroxisome proliferator-activated receptor gamma ($PPAR{\gamma}$), leading to increased glucose transporter 4 (Glut-4) expression and 2-deoxyglucose uptake in differentiated 3T3-L1 cells. Additionally, feeding lignosulfonic acid to diabetic KK-Ay mice suppressed increase of blood glucose level. Conclusion: Lignosulfonic acid may be useful as a functional anti-diabetic component of food.

TonEBP suppresses adipocyte differentiation via modulation of early signaling in 3T3-L1 cells

  • Kim, Soo Jin;Kim, Taehee;Choi, Han Na;Cho, Eun Jung;Park, Jin Bong;Jeon, Byeong Hwa;Lee, Sang Do
    • The Korean Journal of Physiology and Pharmacology
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    • v.20 no.6
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    • pp.649-655
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    • 2016
  • TonEBP belongs to the Rel family of transcription factors and plays important roles in inflammation as well as kidney homeostasis. Recent studies suggest that TonEBP expression is also involved in differentiation of several cell types such as myocytes, chondrocytes, and osteocytes. In this study, we investigated the roles of TonEBP during adipocyte differentiation in 3T3-L1 cells. TonEBP mRNA and protein expression was dramatically reduced during adipocyte differentiation. Sustained expression of TonEBP using an adenovirus suppressed the formation of lipid droplets as well as the expression of FABP4, a marker of differentiated adipocytes. TonEBP also inhibited the expression of $PPAR{\gamma}$, a known master regulator of adipocytes. RNAi-mediated knock down of TonEBP promoted adipocyte differentiation. However, overexpression of TonEBP did not affect adipogenesis after the initiation of differentiation. Furthermore, TonEBP expression suppressed mitotic clonal expansion and insulin signaling, which are required early for adipocyte differentiation of 3T3-L1 cells. These results suggest that TonEBP may be an important regulatory factor in the early phase of adipocyte differentiation.

Inhibitory effect of ethanolic extract of Abeliophyllum distichum leaf on 3T3-L1 adipocyte differentiation

  • Thomas, Shalom Sara;Eom, Ji;Sung, Nak-Yun;Kim, Dong-Sub;Cha, Youn-Soo;Kim, Kyung-Ah
    • Nutrition Research and Practice
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    • v.15 no.5
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    • pp.555-567
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    • 2021
  • BACKGROUND/OBJECTIVES: Abeliophyllum distichum is a plant endemic to Korea, containing several beneficial natural compounds. This study investigated the effect of A. distichum leaf extract (ALE) on adipocyte differentiation. MATERIALS/METHODS: The cytotoxic effect of ALE was analyzed using cell viability assay. 3T3-L1 preadipocytes were differentiated using induction media in the presence or absence of ALE. Lipid accumulation was confirmed using Oil Red O staining. The mRNA expression of adipogenic markers was measured using RT-PCR, and the protein expressions of mitogen-activated protein kinase (MAPK) and peroxisome proliferator-activated receptor gamma (PPAR𝛾) were measured using western blot. Cell proliferation was measured by calculating the incorporation of Bromodeoxyuridine (BrdU) into DNA. RESULTS: ALE reduced lipid accumulation in differentiated adipocytes, as indicated by Oil Red O staining and triglyceride assays. Treatment with ALE decreased the gene expression of adipogenic markers such as Ppar𝛾, CCAAT/enhancer binding protein alpha (C/ebp𝛼), lipoprotein lipase, adipocyte protein-2, acetyl-CoA carboxylase, and fatty acid synthase. Also, the protein expression of PPAR𝛄 was reduced by ALE. Treating the cells with ALE at different time points revealed that the inhibitory effect of ALE on adipogenesis is higher in the early period treatment than in the terminal period. Furthermore, ALE inhibited adipocyte differentiation by reducing the early phase of adipogenesis and mitotic clonal expansion. This was indicated by the lower number of cells in the Synthesis phase of the cell cycle (labeled using BrdU assay) and a decrease in the expression of early adipogenic transcription factors such as C/ebp𝛽 and C/ebp𝛿. ALE suppressed the phosphorylation of MAPK, confirming that the effect of ALE was through the suppression of early phase of adipogenesis. CONCLUSIONS: Altogether, the results of the present study revealed that ALE inhibits lipid accumulation and may be a potential agent for managing obesity.

Psidium guajava L. leaf extract inhibits adipocyte differentiation and improves insulin sensitivity in 3T3-L1 cells

  • Choi, Esther;Baek, Seoyoung;Baek, Kuanglim;Kim, Hye-Kyeong
    • Nutrition Research and Practice
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    • v.15 no.5
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    • pp.568-578
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    • 2021
  • BACKGROUND/OBJECTIVES: Psidium guajava L. (guava) leaves have been shown to exhibit hypoglycemic and antidiabetic effects in rodents. This study investigated the effects of guava leaf extract on adipogenesis, glucose uptake, and lipolysis of adipocytes to examine whether the antidiabetic properties are mediated through direct effects on adipocytes. MATERIALS/METHODS: 3T3-L1 cells were treated with 25, 50, 100 ㎍/mL of methanol extract from guava leaf extract (GLE) or 0.1% dimethyl sulfoxide as a control. Lipid accumulation was evaluated with Oil Red O Staining and AdipoRed assay. Immunoblotting was performed to measure the expression of adipogenic transcription factors, fatty acid synthase (FAS), and AMP-activated protein kinase (AMPK). Glucose uptake under basal or insulin-stimulated condition was measured using a glucose analog 2-[N-(7-nitrobenz-2-oxa-1,3-diazol-4-yl) amino]-2-deoxy-D-glucose. Lipolysis from fully differentiated adipocytes was measured by free fatty acids release into the culture medium in the presence or absence of epinephrine. RESULTS: Oil Red O staining and AdipoRed assay have shown that GLE treatment reduced lipid accumulation during adipocyte differentiation. Mitotic clonal expansion, an early essential event for adipocyte differentiation, was inhibited by GLE treatment. GLE inhibited the expression of transcription factors involved in adipocyte differentiation, such as peroxisome proliferator-activated receptor 𝛄 (PPAR𝛄), CCAAT/enhancer-binding protein α (C/EBPα), and sterol regulatory element-binding protein-1c (SREBP-1c). FAS expression was also decreased while the phosphorylation of AMPK was increased by GLE treatment. In addition, GLE increased insulin-induced glucose uptake into adipocytes. In lipid-filled mature adipocytes, GLE enhanced epinephrine-induced lipolysis but reduced basal lipolysis dose-dependently. CONCLUSIONS: The results show that GLE inhibits adipogenesis and improves adipocyte function by reducing basal lipolysis and increasing insulin-stimulated glucose uptake in adipocytes, which can be partly associated with antidiabetic effects of guava leaves.

SLC3A2 and SLC7A2 Mediate the Exogenous Putrescine-Induced Adipocyte Differentiation

  • Jin, Eom;Juhyun, Choi;Sung-Suk, Suh;Jong Bae, Seo
    • Molecules and Cells
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    • v.45 no.12
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    • pp.963-975
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    • 2022
  • Exogenous polyamines are able to induce life span and improve glucose homeostasis and insulin sensitivity. However, the effects of exogenous polyamines on adipocyte differentiation and which polyamine transporters mediate them have not been elucidated yet. Here, we identified for the first time that exogenous polyamines can clearly stimulate adipocyte differentiation through polyamine transporters, solute carrier family 3 member A2 (SLC3A2) and SLC7A1. Exogenous polyamines markedly promote 3T3-L1 adipocyte differentiation by increasing the intracellular lipid accumulation and the expression of both adipogenic and lipogenic genes in a concentration-dependent manner. In particular, exogenous putrescine mainly regulates adipocyte differentiation in the early and intermediate stages. Moreover, we have assessed the expression of polyamine transporter genes in 3T3-L1 preadipocytes and adipocytes. Interestingly, the putrescine-induced adipocyte differentiation was found to be significantly suppressed in response to a treatment with a polyamine transporter inhibitor (AMXT-1501). Furthermore, knockdown experiments using siRNA that specifically targeted SLC3A2 or SLC7A2, revealed that both SLC3A2 and SLC7A2 act as important transporters in the cellular importing of exogenous putrescine. Thus, the exogenous putrescine entering the adipocytes via cellular transporters is involved in adipogenesis through a modulation of both the mitotic clonal expansion and the expression of master transcription factors. Taken together, these results suggest that exogenous polyamines (such as putrescine) entering the adipocytes through polyamine transporters, can stimulate adipogenesis.

Anti-obesity effect of EGCG and glucosamine-6-phosphate through decreased expression of genes related to adipogenesis and cell cycle arrest in 3T3-L1 adipocytes (Adipogenesis관련 유전자발현감소와 Cell Cycle Arrest를 통한 EGCG와 Glucosamine-6-Phosphate의 Anti-Obesity 효과)

  • Kim, Kkot Byeol;Jang, Seong hee
    • Journal of Nutrition and Health
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    • v.47 no.1
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    • pp.1-11
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    • 2014
  • Purpose: Several studies have proven that EGCG, the primary green tea catechin, and glucosamine-6-phosphate (PGlc) reduce triglyceride contents in 3T3-L1 adipocytes. The objective of this study is to evaluate the combination effect of EGCG and PGlc on decline of accumulated fat in differentiated 3T3-L1 adipocytes. Methods: EGCG and PGlc were administered for 6 day for differentiation of 3T3-L1 adipocytes. Cell viability was measured using the CCK assay kit. In addition, TG accumulation in culture 3T3-L1 adipocytes was investigated by Oil Red O staining. We examined the expres-sion level of several genes and proteins associated with adipogenesis and lipolysis using real-time RT-PCR and Western blot analysis. A flow cytometer Calibar was used to assess the effect of EGCG and PGluco on cell-cycle progression of differentiating 3T3-L1 cells. Results: Intracelluar lipid accumulation was significantly decreased by combination treatment with EGCG $60{\mu}M$ and PGlc $200{\mu}g/m$ compared with control and EGCG treatment alone. In addition, use of combination treatment resulted in directly decreased expression of $PPAR{\gamma}$, $C/EBP{\alpha}$, and SREBP1. In addition, it inhibited adipocyte differentiation and adipogenesis through downstream regulation of adipogenic target genes such as FAS, ACSL1, and LPL, and the inhibitory action of EGCG and PGlc was found to inhibit the mitotic clonal expansion (MCE) process as evidenced by impaired cell cycle entry into S phase and the S to G2/M phase transition of confluent cells and levels of cell cycle regulating proteins such as cyclin A and CDK2. Conclusion: Combination treatment of EGCG and PGlc inhibited adipocyte differentiation through decreased expression of genes related to adipogenesis and adipogenic and cell cycle arrest in early stage of adipocyte differentiation.