FGF signaling in specification of hESC-derived definitive endoderm
(2010) In Lund University Faculty of Medicine Doctoral Dissertation Series 2010:6.- Abstract
- Diabetes affects around 200 million people worldwide. Curing diabetes would
require the prevention of autoimmune destruction of beta cells and restoration of
the beta cell mass restored either through regeneration or transplantation of the
insulin producing cells. Islet transplantation according to the Edmonton protocol
have been the most promising therapeutic option for Diabetes type I patients,
however, lack of cadaveric donor islets is a major obstacle and new strategies need
to be established. Human embryonic stem cells (hESCs) not only offer an excellent
source for establishment of strategies for future regenerative cell therapies and drug
discovery but also... (More) - Diabetes affects around 200 million people worldwide. Curing diabetes would
require the prevention of autoimmune destruction of beta cells and restoration of
the beta cell mass restored either through regeneration or transplantation of the
insulin producing cells. Islet transplantation according to the Edmonton protocol
have been the most promising therapeutic option for Diabetes type I patients,
however, lack of cadaveric donor islets is a major obstacle and new strategies need
to be established. Human embryonic stem cells (hESCs) not only offer an excellent
source for establishment of strategies for future regenerative cell therapies and drug
discovery but also offers en excellent experimental assay for understanding human
pancreas development.
Pancreas originates from the definitive endoderm (DE), one of the three germ
layers. The path from definitive endoderm to pancreatic progenitor cells and finally
the insulin producing beta cells involves sequential cell-fate decisions characterized
by the expression of multiple transcription factors. Governed by the developmental
biological principles that normally control foregut endoderm and pancreas
specification, numerous multifactor protocols for directing mature foregut-derived
cells from hESCs have been reported. However, to establish less complex and more
robust protocols there is a need to further understand the mechanism of action of
individual growth and differentiation factors in specification of human pluripotent
stem cells towards foregut derived cell lineages.
Here, we describe two studies where we have investigated the role of fibroblast
growth factor (FGF) signaling, specifically FGF2 and FGF4, and Retinoic acid (RA)
in specification of the hESC-derived DE development. Studies in lower vertebrates
have demonstrated that FGF2 acts in a restricted manner primarily patterning the
ventral foregut endoderm into liver and lung, whereas FGF4 exhibits broad anteriorposterior
and left-right patterning activities. Furthermore, whereas FGF2 is not
required for ventral pancreas development an inductive role of FGF2 has been
shown during dorsal pancreas formation. Whether FGF2 and FGF4 play a similar
role during human endoderm development remained however unknown. In contrast,
RA has frequently been employed (together with other growth factors) for directed
differentiation of hESCs to pancreatic endoderm.
In the first study we show that FGF2 in a dosage-dependent manner specifies
hESC-derived DE into different foregut lineages such as liver, lung, pancreatic
and intestinal cells. Furthermore, by dissecting the FGF receptor intracellular
pathway that regulates pancreas specification, we demonstrate for the first time to
our knowledge that induction of PDX1+ pancreatic progenitors in part relies on
8
FGF2-mediated activation of the MAPK signaling pathway. In the second study, we
show that FGF4 alone is not sufficient for induction of foregut endoderm but that
in combination with RA it efficiently induces PDX1+ cells from hESC-derived DE.
Specifically, FGF4 promoted cell survival in the differentiating hESCs. Hence, in
contrast to studies in lower vertebrates we demonstrate that FGF4 neither patterns
hESC-derived DE, nor induces PDX1+ pancreatic progenitors suggesting that FGF4
is not responsible for anterior-posterior patterning of the primitive gut during human
development.. Altogether, these observations suggest a broader gut endodermal
patterning activity of FGF2 that corresponds to what has previously been advocated
for FGF4, implying a functional switch from FGF4 to FGF2 during evolution. Thus,
our results provide new knowledge of how cell fate specification of human DE is
controlled – facts that will be of great value for future regenerative cell therapies.
Finally, we present a method for efficient gene targeting in hESCs, which allows the
monitoring of gene expression in living cells. (Less) - Abstract (Swedish)
- Popular Abstract in Swedish
Diabetes är en snabbt växande folksjukdom som redan har drabbat 200 miljoner
världen runt. Det finns två typer av sjukdomen: typ I samt typ II diabetes. Den
förstnämnda drabbar vanligtvis de unga och beror på en s.k. autoimmun reaktion
där kroppens egna immunförsvar av okänd anledning plötsligt inte känner igen
insulincellerna och börjar attackera dem. Typ II diabetes uppstår däremot pga. en
kombination av insulinresistens, samt otillräcklig sekretion av insulin, och bryter
ut i vuxen ålder. Hälsosam kost och motion är av stor vikt för att förhindra typ
II diabetes. Det finns i dagsläget inget botemedel för diabetes och båda... (More) - Popular Abstract in Swedish
Diabetes är en snabbt växande folksjukdom som redan har drabbat 200 miljoner
världen runt. Det finns två typer av sjukdomen: typ I samt typ II diabetes. Den
förstnämnda drabbar vanligtvis de unga och beror på en s.k. autoimmun reaktion
där kroppens egna immunförsvar av okänd anledning plötsligt inte känner igen
insulincellerna och börjar attackera dem. Typ II diabetes uppstår däremot pga. en
kombination av insulinresistens, samt otillräcklig sekretion av insulin, och bryter
ut i vuxen ålder. Hälsosam kost och motion är av stor vikt för att förhindra typ
II diabetes. Det finns i dagsläget inget botemedel för diabetes och båda typerna
av sjukdomen behandlas med insulin som antingen administreras i tablettform
eller som injektioner. För att bota diabetes måste forskarna dels komma på vad
som utlöser den autoimmuna förstörelsen av insulincellerna, dels kunna komma
på ett sätt att bilda nya insulinceller som kan transplanteras till de drabbade. Den
mest framgångsrika behandlingen av typ I diabetes involverar transplantation av
öar med insulinceller. Det finns studier som visar att patienter som genomgått denna
transplantation har kunnat leva utan insulinbehandling i flera år. Men tyvärr har denna
behandling begränsningar, såsom att det krävs flera donatorer till en transplantation,
då ett stort antal av insulinöar krävs för lyckad behandling, och att det krävs livslång
behandling med immunosuppressiva medel för att de transplanterade cellerna inte
ska stötas bort. Det behövs därmed dels alternativa källor till insulinceller, dels
alternativa behandlingsstrategier.
Humana embryonala stamceller har förmågan att bilda vilka celltyper som helst
i människans kropp. Av denna anledning är de väldigt intressanta vid behandling
av degenerativa sjukdomar såsom diabetes, Alzheimers, Parkinson och MS, där de
skulle kunna ersätta den skadade vävnaden. För att kunna styra stamcellerna mot
specifika öden eller celltyper krävs det dock kunskap om exakt vilka signalvägar
som är av betydelse vid bildandet av de specifika cellerna och hur dessa aktiveras.
Genom olika djurmodeller har forskarna lyckats få en inblick i vilka gener och
stimuli som är av betydelse för pankreasutvecklingen, men det finns fortfarande
många luckor innan vi vet exakt hur en insulincell bildas i människan. Genom att
arbeta med humana embryonala stamceller har forskarna även en möjlighet att lära
sig mer om människan och hennes utveckling.
Många olika signaleringsvägar är involverade i att styra de omogna stamcellerna till
insulinproducerande celler. Idag kan vi genom tillsats av olika tillväxtfaktorer, som
har visat sig ha en betydande roll under pankreasutvecklingen i musmodeller, styra
utvecklingen av de omogna stamcellerna till definitivt endoderm, det grodlager som
ger upphov till pankreas, lever, lunga och tarmarna. Vår fortsatta strategi involverar
att hitta nya signaleringsvägar som är av betydelse för att ombilda dessa celler mot
10
ett förstadium till pankreas.
I denna avhandling presenteras två olika protokoll för att styra stamcellerna mot ett
förstadium till pankreas och insulinceller. I det ena protokollet lyckas vi även få ett
förstadium till lever, tarm och lungceller. Dessa nya kunskaper kommer att kunna
användas i vårt fortsatta arbete för att styra stamcellerna mot insulinproducerande
celler, som i framtiden kan användas för transplantation i diabetespatienter. Slutligen
beskrivs även en metod där vi genom genetisk modifiering av stamcellerna lyckas
introducera en ny gen som gör att stamcellerna fluorescerar grönt vid uttryck av
specifika gener. Dessa nya stamcellslinjer är oerhört viktiga i vårt fortsatta arbete
där vi försöker lära oss mer om hur de olika pankreasspecifika generna regleras. (Less)
Please use this url to cite or link to this publication:
https://lup.lub.lu.se/record/1527477
- author
- Ameri, Jacqueline LU
- supervisor
-
- Henrik Semb LU
- opponent
-
- PhD Grapin-Botton, Anne, Swiss Institute for Experimental Cancer Research (ISREC), École Polytechnique Fédérale de Lausanne (EPFL)
- organization
- publishing date
- 2010
- type
- Thesis
- publication status
- published
- subject
- in
- Lund University Faculty of Medicine Doctoral Dissertation Series
- volume
- 2010:6
- pages
- 91 pages
- publisher
- Stem Cells and Pancreas Development Biology
- defense location
- Segerfalksalen, BMC
- defense date
- 2010-01-29 13:00:00
- ISSN
- 1652-8220
- ISBN
- 978-91-86443-20-7
- language
- English
- LU publication?
- yes
- additional info
- The information about affiliations in this record was updated in December 2015. The record was previously connected to the following departments: Stem Cell and Pancreas Developmental Biology (013212044)
- id
- 39535ae5-4b91-41a0-97f5-2cf8256009a1 (old id 1527477)
- date added to LUP
- 2016-04-01 13:38:18
- date last changed
- 2023-04-18 20:32:24
@phdthesis{39535ae5-4b91-41a0-97f5-2cf8256009a1, abstract = {{Diabetes affects around 200 million people worldwide. Curing diabetes would<br/><br> require the prevention of autoimmune destruction of beta cells and restoration of<br/><br> the beta cell mass restored either through regeneration or transplantation of the<br/><br> insulin producing cells. Islet transplantation according to the Edmonton protocol<br/><br> have been the most promising therapeutic option for Diabetes type I patients,<br/><br> however, lack of cadaveric donor islets is a major obstacle and new strategies need<br/><br> to be established. Human embryonic stem cells (hESCs) not only offer an excellent<br/><br> source for establishment of strategies for future regenerative cell therapies and drug<br/><br> discovery but also offers en excellent experimental assay for understanding human<br/><br> pancreas development.<br/><br> Pancreas originates from the definitive endoderm (DE), one of the three germ<br/><br> layers. The path from definitive endoderm to pancreatic progenitor cells and finally<br/><br> the insulin producing beta cells involves sequential cell-fate decisions characterized<br/><br> by the expression of multiple transcription factors. Governed by the developmental<br/><br> biological principles that normally control foregut endoderm and pancreas<br/><br> specification, numerous multifactor protocols for directing mature foregut-derived<br/><br> cells from hESCs have been reported. However, to establish less complex and more<br/><br> robust protocols there is a need to further understand the mechanism of action of<br/><br> individual growth and differentiation factors in specification of human pluripotent<br/><br> stem cells towards foregut derived cell lineages.<br/><br> Here, we describe two studies where we have investigated the role of fibroblast<br/><br> growth factor (FGF) signaling, specifically FGF2 and FGF4, and Retinoic acid (RA)<br/><br> in specification of the hESC-derived DE development. Studies in lower vertebrates<br/><br> have demonstrated that FGF2 acts in a restricted manner primarily patterning the<br/><br> ventral foregut endoderm into liver and lung, whereas FGF4 exhibits broad anteriorposterior<br/><br> and left-right patterning activities. Furthermore, whereas FGF2 is not<br/><br> required for ventral pancreas development an inductive role of FGF2 has been<br/><br> shown during dorsal pancreas formation. Whether FGF2 and FGF4 play a similar<br/><br> role during human endoderm development remained however unknown. In contrast,<br/><br> RA has frequently been employed (together with other growth factors) for directed<br/><br> differentiation of hESCs to pancreatic endoderm.<br/><br> In the first study we show that FGF2 in a dosage-dependent manner specifies<br/><br> hESC-derived DE into different foregut lineages such as liver, lung, pancreatic<br/><br> and intestinal cells. Furthermore, by dissecting the FGF receptor intracellular<br/><br> pathway that regulates pancreas specification, we demonstrate for the first time to<br/><br> our knowledge that induction of PDX1+ pancreatic progenitors in part relies on<br/><br> 8<br/><br> FGF2-mediated activation of the MAPK signaling pathway. In the second study, we<br/><br> show that FGF4 alone is not sufficient for induction of foregut endoderm but that<br/><br> in combination with RA it efficiently induces PDX1+ cells from hESC-derived DE.<br/><br> Specifically, FGF4 promoted cell survival in the differentiating hESCs. Hence, in<br/><br> contrast to studies in lower vertebrates we demonstrate that FGF4 neither patterns<br/><br> hESC-derived DE, nor induces PDX1+ pancreatic progenitors suggesting that FGF4<br/><br> is not responsible for anterior-posterior patterning of the primitive gut during human<br/><br> development.. Altogether, these observations suggest a broader gut endodermal<br/><br> patterning activity of FGF2 that corresponds to what has previously been advocated<br/><br> for FGF4, implying a functional switch from FGF4 to FGF2 during evolution. Thus,<br/><br> our results provide new knowledge of how cell fate specification of human DE is<br/><br> controlled – facts that will be of great value for future regenerative cell therapies.<br/><br> Finally, we present a method for efficient gene targeting in hESCs, which allows the<br/><br> monitoring of gene expression in living cells.}}, author = {{Ameri, Jacqueline}}, isbn = {{978-91-86443-20-7}}, issn = {{1652-8220}}, language = {{eng}}, publisher = {{Stem Cells and Pancreas Development Biology}}, school = {{Lund University}}, series = {{Lund University Faculty of Medicine Doctoral Dissertation Series}}, title = {{FGF signaling in specification of hESC-derived definitive endoderm}}, url = {{https://lup.lub.lu.se/search/files/3492702/1527867.pdf}}, volume = {{2010:6}}, year = {{2010}}, }