Glucocorticoid Signaling Enhances Expression of Glucose-Sensing Molecules in Immature Pancreatic Beta-Like Cells Derived from Murine Embryonic Stem Cells In Vitro
Pluripotent stem cells may serve as an alternative source of beta-like cells for replacement therapy of type 1 diabetes; however, the beta-like cells generated in many differentiation protocols are immature. The maturation of endogenous beta cells involves an increase in insulin expression starting...
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Veröffentlicht in: | Stem cells and development 2018-07, Vol.27 (13), p.898-909 |
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creator | Ghazalli, Nadiah Wu, Xiaoxing Walker, Stephanie Trieu, Nancy Hsin, Li-Yu Choe, Justin Chen, Chialin Hsu, Jasper LeBon, Jeanne Kozlowski, Mark T. Rawson, Jeffrey Tirrell, David A. Yip, M.L. Richard Ku, Hsun Teresa |
description | Pluripotent stem cells may serve as an alternative source of beta-like cells for replacement therapy of type 1 diabetes; however, the beta-like cells generated in many differentiation protocols are immature. The maturation of endogenous beta cells involves an increase in insulin expression starting in late gestation and a gradual acquisition of the abilities to sense glucose and secrete insulin by week 2 after birth in mice; however, what molecules regulate these maturation processes are incompletely known. In this study, we aim to identify small molecules that affect immature beta cells. A cell-based assay, using pancreatic beta-like cells derived from murine embryonic stem (ES) cells harboring a transgene containing an insulin 1-promoter driven enhanced green fluorescent protein reporter, was used to screen a compound library (NIH Clinical Collection-003). Cortisone, a glucocorticoid, was among five positive hit compounds. Quantitative reverse transcription–polymerase chain reaction analysis revealed that glucocorticoids enhance the gene expression of not only insulin 1 but also glucose transporter-2 (
Glut2
;
Slc2a2
) and glucokinase (
Gck
), two molecules important for glucose sensing. Mifepristone, a pharmacological inhibitor of glucocorticoid receptor (GR) signaling, reduced the effects of glucocorticoids on
Glut2
and
Gck
expression. The effects of glucocorticoids on ES-derived cells were further validated in immature primary islets. Isolated islets from 1-week-old mice had an increased
Glut2
and
Gck
expression in response to a 4-day treatment of exogenous hydrocortisone in vitro. Gene deletion of GR in beta cells using rat insulin 2 promoter-driven Cre crossed with GR
flox/flox
mice resulted in a reduced gene expression of
Glut2
, but not
Gck
, and an abrogation of insulin secretion when islets were incubated in 0.5 mM
d
-glucose and stimulated by 17 mM
d
-glucose in vitro. These results demonstrate that glucocorticoids positively regulate glucose sensors in immature murine beta-like cells. |
doi_str_mv | 10.1089/scd.2017.0160 |
format | Article |
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Glut2
;
Slc2a2
) and glucokinase (
Gck
), two molecules important for glucose sensing. Mifepristone, a pharmacological inhibitor of glucocorticoid receptor (GR) signaling, reduced the effects of glucocorticoids on
Glut2
and
Gck
expression. The effects of glucocorticoids on ES-derived cells were further validated in immature primary islets. Isolated islets from 1-week-old mice had an increased
Glut2
and
Gck
expression in response to a 4-day treatment of exogenous hydrocortisone in vitro. Gene deletion of GR in beta cells using rat insulin 2 promoter-driven Cre crossed with GR
flox/flox
mice resulted in a reduced gene expression of
Glut2
, but not
Gck
, and an abrogation of insulin secretion when islets were incubated in 0.5 mM
d
-glucose and stimulated by 17 mM
d
-glucose in vitro. These results demonstrate that glucocorticoids positively regulate glucose sensors in immature murine beta-like cells.</description><identifier>ISSN: 1547-3287</identifier><identifier>EISSN: 1557-8534</identifier><identifier>DOI: 10.1089/scd.2017.0160</identifier><identifier>PMID: 29717618</identifier><language>eng</language><publisher>United States: Mary Ann Liebert, Inc</publisher><subject>Original Research Reports</subject><ispartof>Stem cells and development, 2018-07, Vol.27 (13), p.898-909</ispartof><rights>2018, Mary Ann Liebert, Inc.</rights><rights>Copyright 2018, Mary Ann Liebert, Inc. 2018</rights><lds50>peer_reviewed</lds50><oa>free_for_read</oa><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c431t-238b20568fb3d89a857acf8342c97bfb227cd5f34ee22e3809fc5f185c47c4923</citedby><cites>FETCH-LOGICAL-c431t-238b20568fb3d89a857acf8342c97bfb227cd5f34ee22e3809fc5f185c47c4923</cites></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><link.rule.ids>230,314,780,784,885,27924,27925</link.rule.ids><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/29717618$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><creatorcontrib>Ghazalli, Nadiah</creatorcontrib><creatorcontrib>Wu, Xiaoxing</creatorcontrib><creatorcontrib>Walker, Stephanie</creatorcontrib><creatorcontrib>Trieu, Nancy</creatorcontrib><creatorcontrib>Hsin, Li-Yu</creatorcontrib><creatorcontrib>Choe, Justin</creatorcontrib><creatorcontrib>Chen, Chialin</creatorcontrib><creatorcontrib>Hsu, Jasper</creatorcontrib><creatorcontrib>LeBon, Jeanne</creatorcontrib><creatorcontrib>Kozlowski, Mark T.</creatorcontrib><creatorcontrib>Rawson, Jeffrey</creatorcontrib><creatorcontrib>Tirrell, David A.</creatorcontrib><creatorcontrib>Yip, M.L. Richard</creatorcontrib><creatorcontrib>Ku, Hsun Teresa</creatorcontrib><title>Glucocorticoid Signaling Enhances Expression of Glucose-Sensing Molecules in Immature Pancreatic Beta-Like Cells Derived from Murine Embryonic Stem Cells In Vitro</title><title>Stem cells and development</title><addtitle>Stem Cells Dev</addtitle><description>Pluripotent stem cells may serve as an alternative source of beta-like cells for replacement therapy of type 1 diabetes; however, the beta-like cells generated in many differentiation protocols are immature. The maturation of endogenous beta cells involves an increase in insulin expression starting in late gestation and a gradual acquisition of the abilities to sense glucose and secrete insulin by week 2 after birth in mice; however, what molecules regulate these maturation processes are incompletely known. In this study, we aim to identify small molecules that affect immature beta cells. A cell-based assay, using pancreatic beta-like cells derived from murine embryonic stem (ES) cells harboring a transgene containing an insulin 1-promoter driven enhanced green fluorescent protein reporter, was used to screen a compound library (NIH Clinical Collection-003). Cortisone, a glucocorticoid, was among five positive hit compounds. Quantitative reverse transcription–polymerase chain reaction analysis revealed that glucocorticoids enhance the gene expression of not only insulin 1 but also glucose transporter-2 (
Glut2
;
Slc2a2
) and glucokinase (
Gck
), two molecules important for glucose sensing. Mifepristone, a pharmacological inhibitor of glucocorticoid receptor (GR) signaling, reduced the effects of glucocorticoids on
Glut2
and
Gck
expression. The effects of glucocorticoids on ES-derived cells were further validated in immature primary islets. Isolated islets from 1-week-old mice had an increased
Glut2
and
Gck
expression in response to a 4-day treatment of exogenous hydrocortisone in vitro. Gene deletion of GR in beta cells using rat insulin 2 promoter-driven Cre crossed with GR
flox/flox
mice resulted in a reduced gene expression of
Glut2
, but not
Gck
, and an abrogation of insulin secretion when islets were incubated in 0.5 mM
d
-glucose and stimulated by 17 mM
d
-glucose in vitro. These results demonstrate that glucocorticoids positively regulate glucose sensors in immature murine beta-like cells.</description><subject>Original Research Reports</subject><issn>1547-3287</issn><issn>1557-8534</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2018</creationdate><recordtype>article</recordtype><recordid>eNqFkU9v0zAcQCMEYmNw5Ip85JLif4mdCxJ0ZVTqBFKBq-U4P3eGxO7sZNq-Dp8UZ-0mOHGyZT8_23pF8ZrgBcGyeZdMt6CYiAUmNX5SnJKqEqWsGH86z7koGZXipHiR0k-MaU0lf16c0EYQURN5Wvy-6CcTTIijM8F1aOt2XvfO79DKX2lvIKHV7T5CSi54FCy65xOUW_Bpxi5DD2bqM-c8Wg-DHqcI6Gs-GkFnKfoIoy437hegJfR9QucQ3Q10yMYwoMspOg9oNbTxLvhMb0cYjuDaox9ujOFl8czqPsGr43hWfP-0-rb8XG6-XKyXHzal4YyMJWWypbiqpW1ZJxstK6GNlYxT04jWtpQK01WWcQBKgUncWFNZIivDheENZWfF-4N3P7UDdAb8GHWv9tENOt6poJ36d8e7K7ULN6rGtKm5yIK3R0EM1xOkUQ0umfwX7SFMSVHMGJM0t8loeUBNDClFsI_XEKzmrip3VXNXNXfN_Ju_3_ZIP4TMADsA87L2vnfQQhz_o_0Dh4yzbA</recordid><startdate>20180701</startdate><enddate>20180701</enddate><creator>Ghazalli, Nadiah</creator><creator>Wu, Xiaoxing</creator><creator>Walker, Stephanie</creator><creator>Trieu, Nancy</creator><creator>Hsin, Li-Yu</creator><creator>Choe, Justin</creator><creator>Chen, Chialin</creator><creator>Hsu, Jasper</creator><creator>LeBon, Jeanne</creator><creator>Kozlowski, Mark T.</creator><creator>Rawson, Jeffrey</creator><creator>Tirrell, David A.</creator><creator>Yip, M.L. 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Richard</creatorcontrib><creatorcontrib>Ku, Hsun Teresa</creatorcontrib><collection>PubMed</collection><collection>CrossRef</collection><collection>MEDLINE - Academic</collection><collection>PubMed Central (Full Participant titles)</collection><jtitle>Stem cells and development</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Ghazalli, Nadiah</au><au>Wu, Xiaoxing</au><au>Walker, Stephanie</au><au>Trieu, Nancy</au><au>Hsin, Li-Yu</au><au>Choe, Justin</au><au>Chen, Chialin</au><au>Hsu, Jasper</au><au>LeBon, Jeanne</au><au>Kozlowski, Mark T.</au><au>Rawson, Jeffrey</au><au>Tirrell, David A.</au><au>Yip, M.L. Richard</au><au>Ku, Hsun Teresa</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Glucocorticoid Signaling Enhances Expression of Glucose-Sensing Molecules in Immature Pancreatic Beta-Like Cells Derived from Murine Embryonic Stem Cells In Vitro</atitle><jtitle>Stem cells and development</jtitle><addtitle>Stem Cells Dev</addtitle><date>2018-07-01</date><risdate>2018</risdate><volume>27</volume><issue>13</issue><spage>898</spage><epage>909</epage><pages>898-909</pages><issn>1547-3287</issn><eissn>1557-8534</eissn><abstract>Pluripotent stem cells may serve as an alternative source of beta-like cells for replacement therapy of type 1 diabetes; however, the beta-like cells generated in many differentiation protocols are immature. The maturation of endogenous beta cells involves an increase in insulin expression starting in late gestation and a gradual acquisition of the abilities to sense glucose and secrete insulin by week 2 after birth in mice; however, what molecules regulate these maturation processes are incompletely known. In this study, we aim to identify small molecules that affect immature beta cells. A cell-based assay, using pancreatic beta-like cells derived from murine embryonic stem (ES) cells harboring a transgene containing an insulin 1-promoter driven enhanced green fluorescent protein reporter, was used to screen a compound library (NIH Clinical Collection-003). Cortisone, a glucocorticoid, was among five positive hit compounds. Quantitative reverse transcription–polymerase chain reaction analysis revealed that glucocorticoids enhance the gene expression of not only insulin 1 but also glucose transporter-2 (
Glut2
;
Slc2a2
) and glucokinase (
Gck
), two molecules important for glucose sensing. Mifepristone, a pharmacological inhibitor of glucocorticoid receptor (GR) signaling, reduced the effects of glucocorticoids on
Glut2
and
Gck
expression. The effects of glucocorticoids on ES-derived cells were further validated in immature primary islets. Isolated islets from 1-week-old mice had an increased
Glut2
and
Gck
expression in response to a 4-day treatment of exogenous hydrocortisone in vitro. Gene deletion of GR in beta cells using rat insulin 2 promoter-driven Cre crossed with GR
flox/flox
mice resulted in a reduced gene expression of
Glut2
, but not
Gck
, and an abrogation of insulin secretion when islets were incubated in 0.5 mM
d
-glucose and stimulated by 17 mM
d
-glucose in vitro. These results demonstrate that glucocorticoids positively regulate glucose sensors in immature murine beta-like cells.</abstract><cop>United States</cop><pub>Mary Ann Liebert, Inc</pub><pmid>29717618</pmid><doi>10.1089/scd.2017.0160</doi><tpages>12</tpages><oa>free_for_read</oa></addata></record> |
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title | Glucocorticoid Signaling Enhances Expression of Glucose-Sensing Molecules in Immature Pancreatic Beta-Like Cells Derived from Murine Embryonic Stem Cells In Vitro |
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