Palmitic Acid Methyl Ester Enhances Adipogenic Differentiation in Rat Adipose Tissue-Derived Mesenchymal Stem Cells through a G Protein-Coupled Receptor-Mediated Pathway

Adipogenic differentiation from stem cells has become a research target due to the increasing interest in obesity. It has been indicated that adipocytes can secrete palmitic acid methyl ester (PAME), which is able to regulate stem cell proliferation. However, the effects of PAME on adipogenic differentiation in stem cell remain unclear. Here, we present that the adipogenic differentiation medium supplemented with PAME induced the differentiation of rat adipose tissue-derived mesenchymal stem cells (rAD-MSCs) into adipocyte. rAD-MSCs were treated with PAME for 12 days and then subjected to various analyses. The results from the present study show that PAME significantly increased the levels of adipogenic differentiation markers, PPARγ and Gpd1, and enhanced adipogenic differentiation in rAD-MSCs. Furthermore, the level of GPR40/120 protein increased during induction of adipocyte differentiation in rAD-MSCs. Cotreatment with PAME and a GPR40/120 antagonist together inhibited the PAME-enhanced adipogenic differentiation. Moreover, PAME significantly increased phosphorylation of extracellular signal-regulated kinases (ERK), but not AKT and mTOR. Cotreatment with PAME and a GPR40/120 antagonist together inhibited the PAME-enhanced ERK phosphorylation and adipogenic differentiation. PAME also increased the intracellular Ca2+ levels. Cotreatment with PAME and a Ca2+ chelator or a phospholipase C (PLC) inhibitor prevented the PAME-enhanced ERK phosphorylation and adipogenic differentiation. Our data suggest that PAME activated the GPR40/120/PLC-mediated pathway, which in turn increased the intracellular Ca2+ levels, thereby activating the ERK, and eventually enhanced adipogenic differentiation in rAD-MSCs. The findings from the present study might help get insight into the physiological roles and molecular mechanism of PAME in regulating stem cell differentiation.

UJI FITOKIMIA DAN GC-MS DAUN UROKEP (SENNA ALATA) SEBAGAI BAHAN OBAT PELANGSING TRADISIONAL ASAL NYAPA INDAH, BERAU

Daun Senna alata telah dikenal sebagai obat penyakit kulit akibat parasit. Namun, di Dusun Nyapa Indah, Kabupaten Berau, Kalimantan Timur, daun Senna alata yang dikenal sebagai Urokep, secara tradisional rebusan daun keringnya digunakan sebagai obat pelangsing. Untuk meningkatkan kegunaan herbal menjadi produk obat herbal yang aman dikonsumsi khalayak umum, harus dilakukan proses pengilmiahan melalui uji fitokimia dan GC-MS, demikian pula dengan daun Urokep. Hasil uji fitokimia ekstrak etanol daun Urokep menunjukkan bahwa daun Urokep mengandung tannin, alkaloid, flavonoid, steroid dan triterpenoid. Kandungan tannin dan flavanoid dalam buah atau sayur yang dikonsumsi sehari sekali sudah cukup untuk digunakan sebagai program diet. Hasil uji GC-MS, diduga ekstrak etanol Urokep mengandung dua puluh komponen bahan kimia aktif, diantaranya 18,11% 1,2-Benzenedicarboxylic acid, mono(2-ethylhexyl) ester, 15,7% Palmitic acid, methyl ester dan 7,75% Beta-sitosterol yang dapat digunakan sebagai bahan untuk obat-obatan aktif dalam industri farmasi.Kata kunci : Urokep, Senna alata, pelangsing, fitokimia, GC-MS

Palmitic Acid Methyl Ester Induces G2/M Arrest in Human Bone Marrow-Derived Mesenchymal Stem Cells via the p53/p21 Pathway

Bone marrow-derived mesenchymal cells (BM-MSCs) are able to differentiate into adipocytes, which can secrete adipokines to affect BM-MSC proliferation and differentiation. Recent evidences indicated that adipocytes can secrete fatty acid metabolites, such as palmitic acid methyl ester (PAME), which is able to cause vasorelaxation and exerts anti-inflammatory effects. However, effects of PAME on BM-MSC proliferation remain unclear. The aim of this study was to investigate the effect of PAME on human BM-MSC (hBM-MSC) proliferation and its underlying molecular mechanisms. hBM-MSCs were treated with PAME for 48 h and then subjected to various analyses. The results from the present study show that PAME significantly reduced the levels of G2/M phase regulatory proteins, cyclin-dependent kinase 1 (Cdk1), and cyclin B1 and inhibited proliferation in hBM-MSCs. Moreover, the level of Mdm2 protein decreased, while the levels of p21 and p53 protein increased in the PAME-treated hBM-MSCs. However, PAME treatment did not significantly affect apoptosis/necrosis, ROS generation, and the level of Cdc25C protein. PAME also induced intracellular acidosis and increased intracellular Ca2+ levels. Cotreatment with PAME and Na+/H+ exchanger inhibitors together further reduced the intracellular pH but did not affect the PAME-induced decreases of cell proliferation and increases of the cell population at the G2/M phase. Cotreatment with PAME and a calcium chelator together inhibited the PAME-increased intracellular Ca2+ levels but did not affect the PAME-induced cell proliferation inhibition and G2/M cell cycle arrest. Moreover, the half-life of p53 protein was prolonged in the PAME-treated hBM-MSCs. Taken together, these results suggest that PAME induced p53 stabilization, which in turn increased the levels of p53/p21 proteins and decreased the levels of Cdk1/cyclin B1 proteins, thereby preventing the activation of Cdk1, and eventually caused cell cycle arrest at the G2/M phase. The findings from the present study might help get insight into the physiological roles of PAME in regulating hBM-MSC proliferation.

Triacylglycerol Profile of a Microalga Chlorococcum Sp. as a Potential Biofuel Feedstock

Chlorococcum sp. has been studied as its possibility for biofuel feedstock. This alga was grown in Zarrouk media. After harvesting, lipid extraction was done by pure n-hexane and mixture of chloroform: methanol (2 : 1)) and hexane: ethanol (3 : 1). Highest oil recovery (21.20%) derived from chloroform: methanol (2 : 1) solvent system from dry biomass whereas the lowest (2.83%) came from n-hexane: ethanol solvent system from wet biomass. GC/MS used for the analysis of extracted lipid revealed that, palmitic acid methyl ester (C17H34O2), 9-octadecenoic acid methyl ester (C19H36O2) were major contents of this biofuel. The acquired fatty acid profile indicates that Chlorococcum sp. could be used as promising biofuel feedstock in near future.Journal of Bangladesh Academy of Sciences, Vol. 40, No. 2, 147-153, 2016