Vol 60, No 1 (2022)
Original paper
Published online: 2022-02-25
Candidate genes responsible for lipid droplets formation during adipogenesis simultaneously affect osteoblastogenesis
1
, 1, 1, 1, 1, 1, 1
, 1, 1, 1, 1, 1, 1
Abstract
Introduction. With cellular lipid storage varying, the balance between lipid intake and lipid degradation was a must to keep healthy and determined the level of lipid droplets. Although lipid droplets accumulation had been well demonstrated in adipocytes, gene expression profiling and gene function during adipogenesis and osteoblastogenesis remain unknown.
Material and methods. Here, this work profiled gene transcriptional landscapes of lipid droplets formation during adipogenesis from human mesenchymal stem cells (hMSCs) using RNA-Seq technique. By using RNA interference (RNAi) we investigated the function of candidate genes during adipogenesis and osteoblastogenesis using Oil Red/Alizarin Red/alkaline phosphatase (ALPL) staining and qRT-PCR (quantitative real-time PCR).
Results. Eleven differentially up-regulated genes associated with lipid droplets formation were identified at 3, 5, 7, 14, 21, and 28 days during adipogenesis. Unexpectedly, APOB per se inhibiting adipogenesis weakened osteoblastogenesis and METTL7A facilitating adipogenesis negligibly inhibited osteoblastogenesis according to the phenotypic characterization of adipocytes and osteoblasts and transcriptional condition of biomarkers through lentivirus transfection assays.
Conclusions. The establishment of the gene transcriptional profiling of lipid droplets formation would provide
the molecular switches of hMSCs cell fate determination and the study targets for fat metabolic diseases.
Keywords: human mesenchymal stem cells (hMSCs)adipogenesisosteoblastogenesisAPOBMETTL7ARNA-seqsiRNA
References
- Pittenger MF, Mackay AM, Beck SC, et al. Multilineage potential of adult human mesenchymal stem cells. Science. 1999; 284(5411): 143–147.
- Walther TC, Chung J, Farese RV. Lipid Droplet Biogenesis. Annu Rev Cell Dev Biol. 2017; 33: 491–510.
- Rendina-Ruedy E, Guntur AR, Rosen CJ. Intracellular lipid droplets support osteoblast function. Adipocyte. 2017; 6(3): 250–258.
- Krahmer N, Farese RV, Walther TC. Balancing the fat: lipid droplets and human disease. EMBO Mol Med. 2013; 5(7): 973–983.
- Li L, Zhang H, Yao Y, et al. (-)-Hydroxycitric Acid Suppresses Lipid Droplet Accumulation and Accelerates Energy Metabolism via Activation of the Adiponectin-AMPK Signaling Pathway in Broiler Chickens. J Agric Food Chem. 2019; 67(11): 3188–3197.
- Wang M, Yi X. Bulging and budding of lipid droplets from symmetric and asymmetric membranes: competition between membrane elastic energy and interfacial energy. Soft Matter. 2021; 17(21): 5319–5328.
- Romanauska A, Köhler A. Reprogrammed lipid metabolism protects inner nuclear membrane against unsaturated fat. Dev Cell. 2021; 56(18): 2562–2578.e3.
- Zheng XY, Yu BL, Xie YF, et al. Apolipoprotein A5 regulates intracellular triglyceride metabolism in adipocytes. Mol Med Rep. 2017; 16(5): 6771–6779.
- Casado ME, Huerta L, Marcos-Díaz A, et al. Hormone-sensitive lipase deficiency affects the expression of SR-BI, LDLr, and ABCA1 receptors/transporters involved in cellular cholesterol uptake and efflux and disturbs fertility in mouse testis. Biochim Biophys Acta Mol Cell Biol Lipids. 2021; 1866(12): 159043.
- Pabois O, Ziolek RM, Lorenz CD, et al. Morphology of bile salts micelles and mixed micelles with lipolysis products, from scattering techniques and atomistic simulations. J Colloid Interface Sci. 2021; 587: 522–537.
- Zahradka P, Neumann S, Aukema HM, et al. Adipocyte lipid storage and adipokine production are modulated by lipoxygenase-derived oxylipins generated from 18-carbon fatty acids. Int J Biochem Cell Biol. 2017; 88: 23–30.
- Heid H, Zimbelmann R, Dörflinger Y, et al. Formation and degradation of lipid droplets in human adipocytes and the expression of aldehyde oxidase (AOX). Cell Tissue Res. 2020; 379(1): 45–62.
- Yi X, Liu J, Wu P, et al. The key microRNA on lipid droplet formation during adipogenesis from human mesenchymal stem cells. J Cell Physiol. 2020; 235(1): 328–338.
- Xu X, Li X, Yan R, et al. Gene expression profiling of human bone marrow-derived mesenchymal stem cells during adipogenesis. Folia Histochem Cytobiol. 2016; 54(1): 14–24.
- Yi X, Wu P, Liu J, et al. Identification of the potential key genes for adipogenesis from human mesenchymal stem cells by RNA-Seq. J Cell Physiol. 2019; 234(11): 20217–20227.
- Casado-Díaz A, Santiago-Mora R, Jiménez R, et al. Cryopreserved human bone marrow mononuclear cells as a source of mesenchymal stromal cells: application in osteoporosis research. Cytotherapy. 2008; 10(5): 460–468.
- Nakamura T, Shiojima S, Hirai Y, et al. Temporal gene expression changes during adipogenesis in human mesenchymal stem cells. Biochem Biophys Res Commun. 2003; 303(1): 306–312.
- Wang YL, Lin SP, Hsieh PCH, et al. Concomitant beige adipocyte differentiation upon induction of mesenchymal stem cells into brown adipocytes. Biochem Biophys Res Commun. 2016; 478(2): 689–695.
- Ross SE, Hemati N, Longo KA, et al. Inhibition of adipogenesis by Wnt signaling. Science. 2000; 289(5481): 950–953.
- Donzelli E, Lucchini C, Ballarini E, et al. ERK1 and ERK2 are involved in recruitment and maturation of human mesenchymal stem cells induced to adipogenic differentiation. J Mol Cell Biol. 2011; 3(2): 123–131.
- Chen J, Liu M, Luo X, et al. Exosomal miRNA-486-5p derived from rheumatoid arthritis fibroblast-like synoviocytes induces osteoblast differentiation through the Tob1/BMP/Smad pathway. Biomater Sci. 2020; 8(12): 3430–3442.
- Shi Y, Chen J, Karner CM, et al. Hedgehog signaling activates a positive feedback mechanism involving insulin-like growth factors to induce osteoblast differentiation. Proc Natl Acad Sci U S A. 2015; 112(15): 4678–4683.
- Shi Yu, Fu Y, Tong W, et al. Uniaxial mechanical tension promoted osteogenic differentiation of rat tendon-derived stem cells (rTDSCs) via the Wnt5a-RhoA pathway. J Cell Biochem. 2012; 113(10): 3133–3142.
- Yi X, Wu P, Liu J, et al. Candidate kinases for adipogenesis and osteoblastogenesis from human bone marrow mesenchymal stem cells. Mol Omics. 2021; 17(5): 790–795.
- Anders S, Huber W. Differential expression analysis for sequence count data. Genome Biol. 2010; 11(10): R106.
- Anders S, Pyl PT, Huber W. HTSeq--a Python framework to work with high-throughput sequencing data. Bioinformatics. 2015; 31(2): 166–169.
- Livak KJ, Schmittgen TD. Analysis of relative gene expression data using real-time quantitative PCR and the 2(-Delta Delta C(T)) Method. Methods. 2001; 25(4): 402–408.
- Ansaloni S, Lelkes N, Snyder J, et al. A streamlined sub-cloning procedure to transfer shRNA from a pSM2 vector to a pGIPZ lentiviral vector. J RNAi Gene Silencing. 2010; 6(2): 411–415.
- Gonçalves MA, Janssen JM, Holkers M, et al. Rapid and sensitive lentivirus vector-based conditional gene expression assay to monitor and quantify cell fusion activity. PLoS One. 2010; 5(6): e10954.
- Assis-Ribas T, Forni MF, Winnischofer SM, et al. Extracellular matrix dynamics during mesenchymal stem cells differentiation. Dev Biol. 2018; 437(2): 63–74.
- Wang Z, Chai C, Wang R, et al. Single-cell transcriptome atlas of human mesenchymal stem cells exploring cellular heterogeneity. Clin Transl Med. 2021; 11(12): e650.
- Frith J, Genever P. Transcriptional Control of Mesenchymal Stem Cell Differentiation. Transfus Med Hemother. 2008; 35(3): 216–227.
- van Herpen NA, Schrauwen-Hinderling VB. Lipid accumulation in non-adipose tissue and lipotoxicity. Physiol Behav. 2008; 94(2): 231–241.
- Singh R, Kaushik S, Wang Y, et al. Autophagy regulates lipid metabolism. Nature. 2009; 458(7242): 1131–1135.
- Hirsch J, Batchelor B. Adipose tissue cellularity in human obesity. Clin Endocrinol Metab. 1976; 5(2): 299–311.
- Abu-Elheiga L, Matzuk MM, Abo-Hashema KA, et al. Continuous fatty acid oxidation and reduced fat storage in mice lacking acetyl-CoA carboxylase 2. Science. 2001; 291(5513): 2613–2616.
- Abu-Elheiga L, Brinkley WR, Zhong L, et al. The subcellular localization of acetyl-CoA carboxylase 2. Proc Natl Acad Sci U S A. 2000; 97(4): 1444–1449.
- Soupene E, Kuypers FA. Mammalian long-chain acyl-CoA synthetases. Exp Biol Med (Maywood). 2008; 233(5): 507–521.
- Sirwi A, Hussain MM. Lipid transfer proteins in the assembly of apoB-containing lipoproteins. J Lipid Res. 2018; 59(7): 1094–1102.
- Ohsaki Y, Cheng J, Suzuki M, et al. Lipid droplets are arrested in the ER membrane by tight binding of lipidated apolipoprotein B-100. J Cell Sci. 2008; 121(Pt 14): 2415–2422.
- Suzuki M, Otsuka T, Ohsaki Y, et al. Derlin-1 and UBXD8 are engaged in dislocation and degradation of lipidated ApoB-100 at lipid droplets. Mol Biol Cell. 2012; 23(5): 800–810.
- Contois JH, McConnell JP, Sethi AA, et al. AACC Lipoproteins and Vascular Diseases Division Working Group on Best Practices. Apolipoprotein B and cardiovascular disease risk: position statement from the AACC Lipoproteins and Vascular Diseases Division Working Group on Best Practices. Clin Chem. 2009; 55(3): 407–419.
- Barbier CE, Lind L, Ahlström H, et al. Apolipoprotein B/A-I ratio related to visceral but not to subcutaneous adipose tissue in elderly Swedes. Atherosclerosis. 2010; 211(2): 656–659.
- Lee Yh, Choi SH, Lee KW, et al. Apolipoprotein B/A1 ratio is associated with free androgen index and visceral adiposity and may be an indicator of metabolic syndrome in male children and adolescents. Clin Endocrinol (Oxf). 2011; 74(5): 579–586.
- Pelletier-Beaumont E, Arsenault BJ, Alméras N, et al. Normalization of visceral adiposity is required to normalize plasma apolipoprotein B levels in response to a healthy eating/physical activity lifestyle modification program in viscerally obese men. Atherosclerosis. 2012; 221(2): 577–582.
- Tanoli T, Yue P, Yablonskiy D, et al. Fatty liver in familial hypobetalipoproteinemia: roles of the APOB defects, intra-abdominal adipose tissue, and insulin sensitivity. J Lipid Res. 2004; 45(5): 941–947.
- Semenkovich CF. Insulin resistance and atherosclerosis. J Clin Invest. 2006; 116(7): 1813–1822.
- Du T, Zhang J, Yuan G, et al. Nontraditional risk factors for cardiovascular disease and visceral adiposity index among different body size phenotypes. Nutr Metab Cardiovasc Dis. 2015; 25(1): 100–107.
- Lamantia V, Bissonnette S, Wassef H, et al. ApoB-lipoproteins and dysfunctional white adipose tissue: Relation to risk factors for type 2 diabetes in humans. J Clin Lipidol. 2017; 11(1): 34–45.e2.
- Capece D, D'Andrea D, Begalli F, et al. Enhanced triacylglycerol catabolism by carboxylesterase 1 promotes aggressive colorectal carcinoma. J Clin Invest. 2021; 131(11).
- Ito M, Nagasawa M, Hara T, et al. Differential roles of CIDEA and CIDEC in insulin-induced anti-apoptosis and lipid droplet formation in human adipocytes. J Lipid Res. 2010; 51(7): 1676–1684.
- Zandbergen F, Mandard S, Escher P, et al. The G0/G1 switch gene 2 is a novel PPAR target gene. Biochem J. 2005; 392(Pt 2): 313–324.
- Yang X, Lu X, Lombès M, et al. The G(0)/G(1) switch gene 2 regulates adipose lipolysis through association with adipose triglyceride lipase. Cell Metab. 2010; 11(3): 194–205.
- Baburski AZ, Sokanovic SJ, Bjelic MM, et al. Circadian rhythm of the Leydig cells endocrine function is attenuated during aging. Exp Gerontol. 2016; 73: 5–13.
- Cole NB, Murphy DD, Grider T, et al. Lipid droplet binding and oligomerization properties of the Parkinson's disease protein alpha-synuclein. J Biol Chem. 2002; 277(8): 6344–6352.
- Ohsaki Y, Cheng J, Fujita A, et al. Cytoplasmic lipid droplets are sites of convergence of proteasomal and autophagic degradation of apolipoprotein B. Mol Biol Cell. 2006; 17(6): 2674–2683.
- Zhang H, Wang Y, Li J, et al. Proteome of skeletal muscle lipid droplet reveals association with mitochondria and apolipoprotein a-I. J Proteome Res. 2011; 10(10): 4757–4768.
- Zehmer JK, Bartz R, Bisel B, et al. Targeting sequences of UBXD8 and AAM-B reveal that the ER has a direct role in the emergence and regression of lipid droplets. J Cell Sci. 2009; 122(Pt 20): 3694–3702.
- Lyu Y, Su X, Deng J, et al. Defective differentiation of adipose precursor cells from lipodystrophic mice lacking perilipin 1. PLoS One. 2015; 10(2): e0117536.
- Čopič A, Antoine-Bally S, Giménez-Andrés M, et al. A giant amphipathic helix from a perilipin that is adapted for coating lipid droplets. Nat Commun. 2018; 9(1): 1332.
- Umlauf E, Csaszar E, Moertelmaier M, et al. Association of stomatin with lipid bodies. J Biol Chem. 2004; 279(22): 23699–23709.
- Justesen J, Stenderup K, Ebbesen EN, et al. Adipocyte tissue volume in bone marrow is increased with aging and in patients with osteoporosis. Biogerontology . 2001; 2(3): 165–171.
- Verma S, Rajaratnam JH, Denton J, et al. Adipocytic proportion of bone marrow is inversely related to bone formation in osteoporosis. J Clin Pathol. 2002; 55(9): 693–698.
- Song L, Tuan RS. Transdifferentiation potential of human mesenchymal stem cells derived from bone marrow. FASEB J. 2004; 18(9): 980–982.
- Alexanian AR. Epigenetic modulators promote mesenchymal stem cell phenotype switches. Int J Biochem Cell Biol. 2015; 64: 190–194.
- Naskar S, Kumaran V, Markandeya YS, et al. Neurogenesis-on-Chip: Electric field modulated transdifferentiation of human mesenchymal stem cell and mouse muscle precursor cell coculture. Biomaterials. 2020; 226: 119522.
- Zhang C, Li L, Jiang Y, et al. Space microgravity drives transdifferentiation of human bone marrow-derived mesenchymal stem cells from osteogenesis to adipogenesis. FASEB J. 2018; 32(8): 4444–4458.
- Costa V, Carina V, Raimondi L, et al. MiR-33a Controls hMSCS Osteoblast Commitment Modulating the Yap/Taz Expression Through EGFR Signaling Regulation. Cells. 2019; 8(12).
- Zhang Y, Ling L, Ajay D/O Ajayakumar A, et al. FGFR2 accommodates osteogenic cell fate determination in human mesenchymal stem cells. Gene. 2022 [Epub ahead of print]; 818: 146199.
- Knani L, Bartolini D, Kechiche S, et al. Melatonin prevents cadmium-induced bone damage: First evidence on an improved osteogenic/adipogenic differentiation balance of mesenchymal stem cells as underlying mechanism. J Pineal Res. 2019; 67(3): e12597.
- Li H, Zhou W, Sun S, et al. Microfibrillar-associated protein 5 regulates osteogenic differentiation by modulating the Wnt/β-catenin and AMPK signaling pathways. Mol Med. 2021; 27(1): 153.
