Haseeb Khatri

Introduction: Insulin secreted by pancreatic β cells play a large role in maintaining glucose homeostasis.¹ Type I diabetes mellitus (T1DM) is a chronic autoimmune disease that impacts this homeostasis by destroying β cells, leading to hyperglycemia.² Uncontrolled blood glucose levels can lead to multi-organ dysfunction.¹ Current treatments include the use of exogenous insulin, which has a major shortfall in the inability to maintain homeostasis of blood glucose levels.³ Pancreas and/or islet cell transplantation success is limited by immune rejection, need for use of immunosuppressants, finding a donor match, surgery-related complications, and potential for autoimmune recurrence. Current research is focused on developing stem cell therapy to cure T1DM.² Developing functional β cells in vitro that respond to environmental cues is a barrier to widespread therapy. Based on a 2013 article by Xie et al, only in vivo differentiated β cells responded to environmental cues.⁴ Gene expression can be altered via environmental cues, resulting in changed chromatin structure and gene transcription. The open chromatin structure of human embryonic stem cells (heSC) conveys the pluripotent characteristic in these cells.⁵ For pancreatic differentiation from heSC, the trimethylation of histone H3 at lysines 4 and 27, H3K4me3 and H3K27me3 respectively, play a large role in achieving in vitro differentation.⁴ The PRC-2 Complex accomplishes this methylation; it is balanced by Jumonji and UTX/KDM6A demethylase activity.⁶ Methods: heSCs are taken stepwise through a differentiation pathway as follows: mesendoderm (ME), definitive endoderm (DE), primitve gut tube (GT), posterior foregut (FG), pancreatic endoderm (PE), and functional endocrine cell (FE).^4,6  Low concentrations of both Activin A and Wnt3a induces ME differentiation from heSCs. Activin/Nodal signaling mediators, SMAD2/3, recruit Jumonji to T and WNT3 genes, removing the repressive H3K27me3. The second step is transcription activation via addition of Wnt3a causing the release of β-catenin, which propels the cells into DE differentiation.⁶ A stepwise addition of DETA-NO, valproic acid, and P300i results in FE from PE.⁷ Results: The addition of various signaling molecules and inhibitors altered the temporal expression of H3K27me3, which suppressed the FE activity. The above methods provide a means of producing FE cells that respond to environmental cues in vitro. Discussion: Studies have found that in vitro differentiated stem cells would produce multiple hormones, regardless of environmental cues. Novel methods demonstrate that FE cells can be differentiated in vitro, which may contribute to finding a cure for T1DM.

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2. ThakkarUG,VanikarAV,TrivediHL.Shouldwepracticestemcelltherapyfortype1 diabetes mellitus as precision medicine? Cytotherapy. 2017. http://www.sciencedirect.com.ezproxy.library.tamu.edu/science/article/pii/S14653249 17300440?via%3Dihub#publication-aip-note. Accessed March 23, 2017.
3. Rezania A, Bruin JE, Arora P, et al. Reversal of diabetes with insulin-producing cells derived in vitro from human pluripotent stem cells. Nat Biotechnol 32(11): 1121-1133. http://www.nature.com.ezproxy.library.tamu.edu/nbt/journal/v32/n11/pdf/nbt.3033.pdf. Accessed April 24, 2017.
4. Xie R, Everett L, Lim HW, et al. Dynamic Chromatin Remodeling Mediated by Polycomb Proteins Orchestrates Pancreatic Differentiation of Human Embryonic Stem Cells. Cell Stem Cell 2013; 12: 224-237. http://ac.els-cdn.com.ezproxy.library.tamu.edu/ S1934590912007060/1-s2.0-S1934590912007060-main.pdf?_tid=ea69d812-26c5-11e7- adf4-00000aacb35e&acdnat=1492801894_10f3eee9ede8092d05156f61339993f8. Accessed March 20, 2017.
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6. Wang L, Xu X, Cao Y, et al. Activin/Smad2-induced Histone H3 Lys-27 Trimethylation (H3K27me3) Reduction Is Crucial to Initiate Mesendoderm Differentiation of Human Embryonic Stem Cells. JBC 2016; 292(4): 1339-1350. http://www.jbc.org.ezproxy.library.tamu.edu/content/292/4/1339.full.pdf. Accessed April 21, 2017.
7. Salguero-Aranda C, Tapia-Limonchi R, Cahuana GM, et al. Differentiation of Mouse Embryonic Stem Cells Toward Functional Pancreatic β-Cell Surrogates Through Epigenetic Regulation of Pdx1 by Nitric Oxide. Cell Transplantation 25: 1879-1892. http:// texasamcolstattx.library.ingentaconnect.com.ezproxy.library.tamu.edu/content/cog/ct/ 2016/00000025/00000010/art00011;jsessionid=3j4dqt30uq2ak.x-ic-live-03. Accessed April 21, 2017.