Abstract
Preadipocytes are widely used as an in vitro model to investigate proliferation, adipogenic differentiation, and lipodystrophy; however, cellular physiology and biology are not fully understood in human preadipocytes. The present study was to investigate the expression of transient receptor potential (TRP) channels in human preadipocytes and their potential roles in regulating proliferation and adipogenic differentiation using approaches of confocal microscopy, whole-cell patch voltage-clamp, reverse transcription polymerase chain reaction, Western blot, etc. We found that TRPV2, TRPV4, and TRPM7 channels were abundantly expressed in human preadipocytes. The intracellular Ca2+ transient activated by the TRPV2 activator probenecid was reversed or prevented by ruthenium red, a TRPV2 blocker. The TRPV4 channel activator, 4α-phorbol 12-13-dicaprinate, enhanced intracellular Ca2+ oscillations, and the effect was inhibited by the TRPV4 blocker RN-1734. TRPM7 current was recorded with dialysis of Mg2+-free pipette solution, which was inhibited by the TRP channel blocker 2-aminoethoxydiphenyl borate and enhanced by acidic extracellular pH. Silencing TRPV2 or TRPM7, but not TRPV4, significantly reduced cell proliferation via inhibiting cyclin D1, cyclin E, and p-ERK1/2. Interestingly, individually silencing these three channels decreased adipogenic differentiation of human preadipocytes by reducing p-Akt kinase. Our results demonstrate for the first time that functional TRPV2, TRPV4, and TRPM7 channels are abundantly expressed in human preadipocytes. TRPV2 and TRPM7, but not TRPV4, regulate cell proliferation via activating cyclin D1, cyclin E, and p-ERK1/2, while they are all involved in adipogenesis in human preadipocytes via phosphorylating Akt kinase.
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References
Adams M, Montague CT, Prins JB, Holder JC, Smith SA, Sanders L, Digby JE, Sewter CP, Lazar MA, Chatterjee VK, O'Rahilly S (1997) Activators of peroxisome proliferator-activated receptor gamma have depot-specific effects on human preadipocyte differentiation. J Clin Invest 100:3149–3153. doi:10.1172/jci119870
Aoyagi K, Ohara-Imaizumi M, Nishiwaki C, Nakamichi Y, Ueki K, Kadowaki T, Nagamatsu S (2012) Acute inhibition of PI3K-PDK1-Akt pathway potentiates insulin secretion through upregulation of newcomer granule fusions in pancreatic beta-cells. PloS one 7:e47381. doi:10.1371/journal.pone.0047381
Armani A, Mammi C, Marzolla V, Calanchini M, Antelmi A, Rosano GM, Fabbri A, Caprio M (2010) Cellular models for understanding adipogenesis, adipose dysfunction, and obesity. J Cell Biochem 110:564–572. doi:10.1002/jcb.22598
Baldoli E, Maier JA (2012) Silencing TRPM7 mimics the effects of magnesium deficiency in human microvascular endothelial cells. Angiogenesis 15:47–57. doi:10.1007/s10456-011-9242-0
Bang S, Kim KY, Yoo S, Lee SH, Hwang SW (2007) Transient receptor potential V2 expressed in sensory neurons is activated by probenecid. Neurosci Lett 425:120–125. doi:10.1016/j.neulet.2007.08.035
Bates-Withers C, Sah R, Clapham DE (2011) TRPM7, the Mg(2+) inhibited channel and kinase. Adv Exp Med Biol 704:173–183. doi:10.1007/978-94-007-0265-3_9
Cheng H, Beck A, Launay P, Gross SA, Stokes AJ, Kinet JP, Fleig A, Penner R (2007) TRPM4 controls insulin secretion in pancreatic beta-cells. Cell calcium 41:51–61. doi:10.1016/j.ceca.2006.04.032
Clapham DE (2003) TRP channels as cellular sensors. Nature 426:517–524. doi:10.1038/nature02196
Dadon D, Minke B (2010) Cellular functions of transient receptor potential channels. Int J Biochem Cell Biol 42:1430–1445. doi:10.1016/j.biocel.2010.04.006
Damann N, Voets T, Nilius B (2008) TRPs in our senses. Curr Biol 18(18):R880–889. doi:10.1016/j.cub.2008.07.063
Deng XL, Lau CP, Lai K, Cheung KF, Lau GK, Li GR (2007) Cell cycle-dependent expression of potassium channels and cell proliferation in rat mesenchymal stem cells from bone marrow. Cell Prolif 40:656–670. doi:10.1111/j.1365-2184.2007.00458.x
Dhaka A, Viswanath V, Patapoutian A (2006) Trp ion channels and temperature sensation. Annu Rev Neurosci 29:135–161. doi:10.1146/annurev.neuro.29.051605.112958
Earley S (2010) Vanilloid and melastatin transient receptor potential channels in vascular smooth muscle. Microcirculation 17:237–249. doi:10.1111/j.1549-8719.2010.00026.x
Federico A, D'Aiuto E, Borriello F, Barra G, Gravina AG, Romano M, De Palma R (2010) Fat: a matter of disturbance for the immune system. World J Gastroenterol 16:4762–4772
Grant AC, Ortiz-Colon G, Doumit ME, Buskirk DD (2008) Optimization of in vitro conditions for bovine subcutaneous and intramuscular preadipocyte differentiation. J Anim Sci 86:73–82. doi:10.2527/jas.2007-0379
Gregoire FM, Smas CM, Sul HS (1998) Understanding adipocyte differentiation. Physiol Rev 78:783–809
Guerre-Millo M (2004) Adipose tissue and adipokines: for better or worse. Diabetes Metab 30:13–19
Hu H, He ML, Tao R, Sun HY, Hu R, Zang WJ, Yuan BX, Lau CP, Tse HF, Li GR (2009) Characterization of ion channels in human preadipocytes. J Cell Physiol 218:427–435. doi:10.1002/jcp.21617
Hu R, He ML, Hu H, Yuan BX, Zang WJ, Lau CP, Tse HF, Li GR (2009) Characterization of calcium signaling pathways in human preadipocytes. J Cell Physiol 220:765–770. doi:10.1002/jcp.21823
Inoue K, Xiong ZG (2009) Silencing TRPM7 promotes growth/proliferation and nitric oxide production of vascular endothelial cells via the ERK pathway. Cardiovasc Res 83:547–557. doi:10.1093/cvr/cvp153
Jiang J, Li M, Yue L (2005) Potentiation of TRPM7 inward currents by protons. J Gen Physiol 126:137–150. doi:10.1085/jgp.200409185
Kajiya H, Okamoto F, Nemoto T, Kimachi K, Toh-Goto K, Nakayana S, Okabe K (2010) RANKL-induced TRPV2 expression regulates osteoclastogenesis via calcium oscillations. Cell Calcium 48:260–269. doi:10.1016/j.ceca.2010.09.010
Kanzaki M, Zhang YQ, Mashima H, Li L, Shibata H, Kojima I (1999) Translocation of a calcium-permeable cation channel induced by insulin-like growth factor-I. Nat Cell Biol 1:165–170. doi:10.1038/11086
Kassmann M, Harteneck C, Zhu Z, Nurnberg B, Tepel M, Gollasch M (2013) Transient receptor potential vanilloid 1 (TRPV1), TRPV4, and the kidney. Acta physiologica (Oxford, England) 207:546–564. doi:10.1111/apha.12051
Koch SE, Gao X, Haar L, Jiang M, Lasko VM, Robbins N, Cai W, Brokamp C, Varma P, Tranter M, Liu Y, Ren X, Lorenz JN, Wang HS, Jones WK, Rubinstein J (2012) Probenecid: novel use as a non-injurious positive inotrope acting via cardiac TRPV2 stimulation. J Mol Cell Cardiol 53:134–144. doi:10.1016/j.yjmcc.2012.04.011
Li GR, Deng XL, Sun H, Chung SS, Tse HF, Lau CP (2006) Ion channels in mesenchymal stem cells from rat bone marrow. Stem Cells 24:1519–1528. doi:10.1634/stemcells.2005-0307
Li GR, Wang HB, Qin GW, Jin MW, Tang Q, Sun HY, Du XL, Deng XL, Zhang XH, Chen JB, Chen L, Xu XH, Cheng LC, Chiu SW, Tse HF, Vanhoutte PM, Lau CP (2008) Acacetin, a natural flavone, selectively inhibits human atrial repolarization potassium currents and prevents atrial fibrillation in dogs. Circulation 117:2449–2457. doi:10.1161/CIRCULATIONAHA.108.769554
Li M, Jiang J, Yue L (2006) Functional characterization of homo- and heteromeric channel kinases TRPM6 and TRPM7. J Gen Physiol 127:525–537. doi:10.1085/jgp.200609502
Liu H, Yang X, Zhang Y, Dighe A, Li X, Cui Q (2012) Fullerol antagonizes dexamethasone-induced oxidative stress and adipogenesis while enhancing osteogenesis in a cloned bone marrow mesenchymal stem cell. J Orthop Res 30:1051–1057. doi:10.1002/jor.22054
Martin E, Dahan D, Cardouat G, Gillibert-Duplantier J, Marthan R, Savineau JP, Ducret T (2012) Involvement of TRPV1 and TRPV4 channels in migration of rat pulmonary arterial smooth muscle cells. Pflugers Archiv 464:261–272. doi:10.1007/s00424-012-1136-5
Mathie A (2010) Ion channels as novel therapeutic targets in the treatment of pain. J Pharm Pharmacol 62:1089–1095. doi:10.1111/j.2042-7158.2010.01131.x
Mergler S, Valtink M, Taetz K, Sahlmuller M, Fels G, Reinach PS, Engelmann K, Pleyer U (2011) Characterization of transient receptor potential vanilloid channel 4 (TRPV4) in human corneal endothelial cells. Exp Eye Res 93:710–719. doi:10.1016/j.exer.2011.09.021
Moffat J, Grueneberg DA, Yang X, Kim SY, Kloepfer AM, Hinkle G, Piqani B, Eisenhaure TM, Luo B, Grenier JK, Carpenter AE, Foo SY, Stewart SA, Stockwell BR, Hacohen N, Hahn WC, Lander ES, Sabatini DM, Root DE (2006) A lentiviral RNAi library for human and mouse genes applied to an arrayed viral high-content screen. Cell 124:1283–1298. doi:10.1016/j.cell.2006.01.040
Monet M, Gkika D, Lehen'kyi V, Pourtier A, Vanden Abeele F, Bidaux G, Juvin V, Rassendren F, Humez S, Prevarsakaya N (2009) Lysophospholipids stimulate prostate cancer cell migration via TRPV2 channel activation. Biochimica et biophysica acta 1793:528–539. doi:10.1016/j.bbamcr.2009.01.003
Montell C (2005) The TRP superfamily of cation channels. Sci STKE 272:re3. doi:10.1126/stke.2722005re3
Morelli MB, Nabissi M, Amantini C, Farfariello V, Ricci-Vitiani L, di Martino S, Pallini R, Larocca LM, Caprodossi S, Santoni M, De Maria R, Santoni G (2012) The transient receptor potential vanilloid-2 cation channel impairs glioblastoma stem-like cell proliferation and promotes differentiation. Int J Cancer 131:E1067–E1077. doi:10.1002/ijc.27588
Motter AL, Ahern GP (2008) TRPV1-null mice are protected from diet-induced obesity. FEBS letters 582:2257–2262. doi:10.1016/j.febslet.2008.05.021
Nadler MJ, Hermosura MC, Inabe K, Perraud AL, Zhu Q, Stokes AJ, Kurosaki T, Kinet JP, Penner R, Scharenberg AM, Fleig A (2001) LTRPC7 is a Mg.ATP-regulated divalent cation channel required for cell viability. Nature 411:590–595. doi:10.1038/35079092
Nilius B (2013) Transient receptor potential TRP channels as therapeutic drug targets: next round! Curr Top Med Chem 13:244–246
Palazzo E, Rossi F, de Novellis V, Maione S (2013) Endogenous modulators of TRP channels. Curr Top Med Chem 13:398–407
Panda S, Nayak SK, Campo B, Walker JR, Hogenesch JB, Jegla T (2005) Illumination of the melanopsin signaling pathway. Science 307:600–604. doi:10.1126/science.1105121
Penner R, Fleig A (2007) The Mg2+ and Mg2+-nucleotide-regulated channel-kinase TRPM7. Handb Exp Pharmacol 179:313–328. doi:10.1007/978-3-540-34891-7_19
Santoni G, Farfariello V, Amantini C (2011) TRPV channels in tumor growth and progression. Adv Exp Med Biol 704:947–967. doi:10.1007/978-94-007-0265-3_49
Schlesinger JB, van Harmelen V, Alberti-Huber CE, Hauner H (2006) Albumin inhibits adipogenesis and stimulates cytokine release from human adipocytes. Am J Physiol Cell Physiol 291:C27–C33. doi:10.1152/ajpcell.00172.2005
Sethi JK, Vidal-Puig AJ (2007) Thematic review series: adipocyte biology. Adipose tissue function and plasticity orchestrate nutritional adaptation. J Lipid Res 48:1253–1262. doi:10.1194/jlr.R700005-JLR200
Shapovalov G, Lehen'kyi V, Skryma R, Prevarskaya N (2011) TRP channels in cell survival and cell death in normal and transformed cells. Cell Calcium 50:295–302. doi:10.1016/j.ceca.2011.05.006
Sonkusare SK, Bonev AD, Ledoux J, Liedtke W, Kotlikoff MI, Heppner TJ, Hill-Eubanks DC, Nelson MT (2012) Elementary Ca2+ signals through endothelial TRPV4 channels regulate vascular function. Science 336:597–601. doi:10.1126/science.1216283
Sukumar P, Sedo A, Li J, Wilson LA, O'Regan D, Lippiat JD, Porter KE, Kearney MT, Ainscough JF, Beech DJ (2012) Constitutively active TRPC channels of adipocytes confer a mechanism for sensing dietary fatty acids and regulating adiponectin. Circ Res 111:191–200. doi:10.1161/CIRCRESAHA.112.270751
Tao R, Lau CP, Tse HF, Li GR (2008) Regulation of cell proliferation by intermediate-conductance Ca2+-activated potassium and volume-sensitive chloride channels in mouse mesenchymal stem cells. Am J Physiol Cell Physiol 295:C1409–C1416. doi:10.1152/ajpcell.00268.2008
Tao R, Sun HY, Lau CP, Tse HF, Lee HC, Li GR (2011) Cyclic ADP ribose is a novel regulator of intracellular Ca2+ oscillations in human bone marrow mesenchymal stem cells. J Cell Mol Med 15:2684–2696. doi:10.1111/j.1582-4934.2011.01263.x
Troidl C, Troidl K, Schierling W, Cai WJ, Nef H, Mollmann H, Kostin S, Schimanski S, Hammer L, Elsasser A, Schmitz-Rixen T, Schaper W (2009) Trpv4 induces collateral vessel growth during regeneration of the arterial circulation. J Cell Mol Med 13:2613–2621. doi:10.1111/j.1582-4934.2008.00579.x
Ueda T, Shikano M, Kamiya T, Joh T, Ugawa S (2011) The TRPV4 channel is a novel regulator of intracellular Ca2+ in human esophageal epithelial cells. Am J Physiol Gastrointest Liver Physiol 301:G138–G147. doi:10.1152/ajpgi.00511.2010
Venkatachalam K, Montell C (2007) TRP channels. Annu Rev Biochem 76:387–417. doi:10.1146/annurev.biochem.75.103004.142819
Vriens J, Appendino G, Nilius B (2009) Pharmacology of vanilloid transient receptor potential cation channels. Mol Pharmacol 75:1262–1279. doi:10.1124/mol.109.055624
Vriens J, Janssens A, Prenen J, Nilius B, Wondergem R (2004) TRPV channels and modulation by hepatocyte growth factor/scatter factor in human hepatoblastoma (HepG2) cells. Cell Calcium 36:19–28. doi:10.1016/j.ceca.2003.11.006
Wang Y, Wang DH (2011) Protective effect of TRPV1 against renal fibrosis via inhibition of TGF-beta/Smad signaling in DOCA-salt hypertension. Mol Med 17:1204–1212. doi:10.2119/molmed.2011.00063
Woudenberg-Vrenken TE, Bindels RJ, Hoenderop JG (2009) The role of transient receptor potential channels in kidney disease. Nat Rev Nephrol 5:441–449. doi:10.1038/nrneph.2009.100
Zhang LL, Yan Liu D, Ma LQ, Luo ZD, Cao TB, Zhong J, Yan ZC, Wang LJ, Zhao ZG, Zhu SJ, Schrader M, Thilo F, Zhu ZM, Tepel M (2007) Activation of transient receptor potential vanilloid type-1 channel prevents adipogenesis and obesity. Circ Res 100:1063–1070. doi:10.1161/01.RES.0000262653.84850.8b
Zhang YH, Sun HY, Chen KH, Du XL, Liu B, Cheng LC, Li X, Jin MW, Li GR (2012) Evidence for functional expression of TRPM7 channels in human atrial myocytes. Basic Res Cardiol 107:282. doi:10.1007/s00395-012-0282-4
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This work was supported in part by Sun Chieh Yeh Heart Foundation of Hong Kong. Hui Che is supported by a postgraduate studentship of University of Hong Kong.
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Che, H., Yue, J., Tse, HF. et al. Functional TRPV and TRPM channels in human preadipocytes. Pflugers Arch - Eur J Physiol 466, 947–959 (2014). https://doi.org/10.1007/s00424-013-1355-4
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DOI: https://doi.org/10.1007/s00424-013-1355-4