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Induction of Pluripotency:
8 C w/ c4 B' N4 z. H' yFrom Mouse to Human
7 n; q: U2 S1 Z' |3 HHolm Zaehres11 l& c- S3 M! B; W2 }# }1 g* N+ }
and Hans R. Schöler1,
% `% ^; ~7 }. V8 Y N*5 h, @/ X- N3 U5 w7 d; B. Y
1$ ~! B& K$ }: k
Max Planck Institute for Molecular Biomedicine, Department of Cell and Developmental Biology, Münster, NRW 48149, Germany
" H' f4 P; y8 D/ s*Correspondence: schoeler@mpi-muenster.mpg.de
: }, H- r( ^( ]( W- E5 R# q4 aDOI 10.1016/j.cell.2007.11.020+ ^5 b% n! C/ S! H: {: y
In this issue of Cell, Takahashi et al. (2007) transfer their seminal work on somatic cell
+ L: E) q5 k* l- w- R( [reprogramming from the mouse to human. By overexpressing the transcription factor ; {0 U" W3 t* N! m6 V4 S
quartet of Oct4, Sox2, Klf4, and c-Myc in adult human fbroblasts, they successfully 2 i n3 }% @8 z! `
isolate human pluripotent stem cells that resemble human embryonic stem cells by all
7 g6 B/ T) s7 v3 e+ nmeasured criteria. This is a signifcant turning point in nuclear reprogramming research
% b5 r- C" ~6 {, {' Qwith broad implications for generating patient-specifc pluripotent stem cells for research
+ U! F- L5 D4 Pand therapeutic applications.1 l, Y8 S+ V3 I
This year’s three Physiology or Medi-
- P/ T! H9 [* A% ^5 x- ycine Nobel Laureates—Martin Evans,
9 f8 A9 W% v+ j& V+ MMario Capecchi, and Oliver Smithies—6 t+ R; o3 z- Z0 K( z
will be honored in Stockholm in 10 , a; s7 s% C: z" \: g
days time for their discovery of DNA
+ y! @+ V% i/ m* @! Q" n( erecombination and the development & Y+ v0 J" S K R" f1 O
of mouse embryonic stem (ES) cell
4 V1 u7 ]2 q f; _3 ~technology. It was Martin Evans who : P! Y5 P- @( A/ e1 p, x9 V
discovered how to make mouse ES
" T, s3 n' P) B0 {8 [: \& @$ Rcells, enabling any genetic alteration
# j7 H1 ]) J% J) m# `. |# qto be transferred to the germline and $ c" |5 u0 s5 t9 Y- F
hence to the next generation (Evans
1 g9 m5 p1 S, q4 Uand Kaufman, 1981; Martin, 1981). $ q6 t5 C; n D' P+ I& ]
Before this breakthrough, researchers
) Q, s: o" g: A7 N, ~: gstudied mouse embryonal carcinoma
! [* h* f4 }4 I* e4 w/ B7 Jcells derived from tumors, which
- m' g ^" c' h# G$ a; ?could form every mouse cell lineage 4 x- F7 m! ^& a
except the germline. Combining DNA , M N8 ~3 A) {+ r
recombination and mouse ES cell 1 z& X# t2 l+ k# A7 m; M9 a" j
technology revolutionized an entire 0 Z2 B" k3 \4 ^4 t) p, A, h. }
feld of research, forming the basis for
0 s7 B9 e7 j9 L- c$ W, ~4 P3 Q& wstudying and understanding the roles
7 ?: R$ y1 `" j% Z, Q8 Z$ Cof numerous genes in embryonic / ?+ ~' P; T# i/ t6 X5 i
development, adult physiology, dis-
+ @/ y# X7 [5 E& E T7 q& wease, and aging. To date, more than
. H# ~1 b+ r. W9 \$ P3 d+ C500 mouse models of human disor-: q! u' y1 v) J: {) E
ders have been generated. Now, with
7 ]4 C) Q9 ]& B9 q# Z$ B: gthe study by Takahashi et al. (2007)
9 }- C% M, g0 T5 g: `2 Apublished in this issue of Cell, another
/ d2 `( q3 ]) ?3 l8 l) q# Qimportant revolution is taking place.
7 E% j/ Y( B+ kLast summer, Takahashi and
+ Q( E. ~% G/ H% v/ r" ~Yamanaka (2006) stunned the scientifc * k6 y; @7 p7 j" o% A! j
community with their study showing 4 D& H I- V9 V
molecular reprogramming of mouse
/ E9 `3 E- Z& A1 s; `; `- Hsomatic cells into induced pluripotent ' M: g, Y+ K5 c8 @1 R- n7 ^/ Q
stem (iPS) cells using just four factors: 3 F1 q6 r1 V' [$ k( c+ x
Oct4, Sox2, Klf4, and c-Myc. Their 6 @" f+ u- s( D; U; H
elegant but demanding approach of W/ D$ q9 \. `' e# g* d
screening for a cocktail of factors that
; S) s1 V7 _9 B B9 R# p$ wcould reprogram mouse fbroblasts * P9 s+ k2 U+ D( p$ W/ k. }" N
starting from 24 candidate genes paid 8 ?9 W5 S, z2 `' r9 c4 d7 v. o G
off with their detailed description of iPS 1 I$ x9 I7 }9 c& P9 X& A
cells, which are almost indistinguish-' h- R- w' K% y0 A* \
able from mouse ES cells. As with all
6 t" A+ }- `- Z8 _4 [2 Y5 k( B1 v a( Hscientifc discoveries, these exciting + D8 @/ s) l: w+ x* k* Z+ G
fndings had to be reproduced. Sev-
* t* p. K' r2 G& }) m# f. ]8 ^eral studies published this year not ) v2 v* a) g# Z- L
only reproduced but also extended
: Y' q% O* \" @the Takahashi and Yamanaka fndings ' l' r' k% P* t* M
by demonstrating the pluripotency and
% R' ?, T1 Z5 [differentiation potential of mouse iPS
9 O! o% y6 \3 d4 h- U. R: x1 ?cells in rigorous developmental assays
6 `8 Z6 z. [$ j2 @" e(Maherali et al., 2007; Okita et al., 2007;
9 u! L! E$ W5 qWernig et al., 2007).: |1 U: @% j% y2 _ r
In their new study, Takahashi,
/ H" D5 V1 `( N7 r+ Z% FYamanaka, and their colleagues
: N6 k Z# E: H; P(Takahashi et al., 2007) now translate 5 O+ o" Z2 {" _: s
their remarkable fndings from mouse
; n% M- ]( Z( F1 z& W4 Zto human (see Figure 1). They selected
0 z' P- g: m& x z iadult human dermal fbroblasts and
- A- \ ^& G3 Btwo other human fbroblast popula-
3 Q6 s# o; F& @; f( d2 `1 x. Mtions (from synovial tissue and neo-
5 p. m1 o8 ], ynatal foreskin) from different human 1 r' M* n0 ]; F) A
donors as their reprogramming target 5 e7 y( T0 T! \" r1 r7 M% W8 D! a
cell populations. They then trans-( a+ u+ Y: j% D* K) D/ j! I
duced the human fbroblast cultures 4 J( _. b% c9 u" k; h5 _6 R3 k! `
with retroviral vectors carrying trans-
6 ]* F2 m! h) C* rgenes for the human versions of Oct4, 2 d u7 J# ~" U- `
Sox2, Klf4, and c-Myc and cultured " R7 e/ X3 R( ]- \
the cells under human ES cell culture ( _1 W( U1 P4 t# f0 i* Q; O' P, B
conditions. Thirty days after transduc-
2 B5 k* f8 p3 z; K8 f$ O/ Gtion, the culture plates were covered
3 f/ a; h( U& D4 n# o9 Hwith human ES cell-like iPS colonies $ x/ ?& N' {- J- e. m: w
(among other colonies), which could
# F2 @. Y. h" e5 Z9 f; e$ H: hbe further propagated and expanded. 2 \3 S5 g7 U" A- K' r; ^
The retroviral vectors enabled silenc-1 j1 w' x( a4 {
ing of all four transgenes after human 6 O9 G/ [1 E0 `9 U1 J8 Z
iPS formation (as found in the mouse
3 E; @, T: _" k0 ysystem) indicating that the iPS cells
3 q& M3 Y+ f" n) m1 d% X( l3 T: vare fully reprogrammed and no longer 4 ?$ {/ ^; K1 G+ a
depend on transgene expression.
* P3 u) ?9 s, i/ `Unlike the mouse study, human
, w/ ~. Y s5 J+ eiPS cells were generated without any 2 U, G7 k" |4 c" |& d8 W* E X
genetic selection procedures. Given
1 G8 r/ `5 c" z$ a( n( `4 Jthe lower mitotic index of human ES
4 c3 D5 i5 E; p- ?6 zcells, it is not surprising that the gen-
7 U/ }% x. {% r7 `3 J6 Y$ }eration of human iPS cells takes nota-4 W6 v( s! r0 s) ~3 m8 u& z
bly longer than in the mouse system.
) f# L+ V$ T" Z7 xThe authors subjected their human 0 Y5 u3 L F0 F% E: j8 w
iPS cells to a panel of assays to com-
6 A* E3 Q2 V! s" T j/ P) ipare them with human ES cells. These - D5 K: x+ q+ j: m& _
assays included morphological stud-
( g/ H: _3 ^) g' Y, t: N) w* p% pies, surface-marker expression, epi-
0 K4 t5 |. s* O, `genetic status, formation of embryoid 7 {6 o+ q1 o, j6 _/ z2 T3 O
bodies in vitro, directed differentia-
8 ^+ j+ ~7 K( ytion into neural cells and beating car-
$ q- ^5 B4 L6 L8 @* j( I% W9 l9 Qdiomyocytes (according to human
' O l, y- Z5 c- N G% ?( g9 G2 dES cell differentiation protocols), and 6 D' d* A& G ]* {7 V
fnally teratoma formation in vivo. 9 I. C: m, J- r8 P
DNA microarray analysis revealed + ?3 _. y- ^6 o
the remarkable degree of similar- r# r$ H5 A' R/ ?
ity between the global gene expres-
5 k8 h/ ~$ `: N( [- F" g7 P+ h' M# usion patterns of human iPS cells and
+ M% Z+ J5 Z5 e. J$ O- o) Bhuman ES cells. Notably, genomic
/ b4 x- j; b- B- c$ z1 RDNA analysis as well as analysis of 4 {: ]* ?) V8 ?' O. n5 n2 H
short tandem repeats demonstrated
$ M& P. a, [. N7 g1 fthe genetic origin of independent 5 [# y' @8 x: S* T
human iPS clones from their parental
, g* h6 l" W% {fbroblast populations.
3 f* ~1 ?, M1 |+ HThe derivation of mouse and then # s X- _& h8 J8 x
human ES cells (Thomson et al., 1998)
4 f: l. c% q, Aas the gold standard of pluripotent
! y# l% F% p/ I" Estem cell populations has necessarily 7 E2 l; n4 }/ D
led to emphasis on differences in the % {! Z6 J$ z5 b' L: Y* L
regulation of self-renewal between ; v( I% V* m3 k$ }9 P; L |
mouse and human ES cells. For ! q# D% E' d- B. b
example, human ES cells depend on 9 n% D" S; D" i2 |/ D
bFGF for self-renewal, whereas their
' K8 W, x+ c7 pmouse counterparts depend on the
' l6 W$ A4 E' JLif/Stat3 pathway; BMP is involved in
G) L; a2 e9 V. imouse ES cell self-renewal, whereas $ o, ]! U& }: h9 i
in human ES cells it induces differen-: E6 k( ?' M, }! H# a
tiation. Extrinsic factors and signals 5 p [9 I. i% S: C
for maintaining pluripotency may dif-
/ ~/ J- C% n% d# {fer between mouse and human. How-
4 F4 {4 @; L- Rever, the ability to translate somatic : M" c4 F/ i6 V* Y9 U6 C
cell reprogramming from mouse to p! {" T$ F# I1 g
human using the same transcription
# I. ]7 S, n' j5 p- vfactor quartet further emphasizes the
' w" Y+ h4 H! J0 _7 x6 M" M: M. Fconserved nature of the Oct4/Sox2
, z$ T/ a/ y, |; O( g9 Etranscription factor network that
B& J8 Y! P9 Dcontrols self-renewal of mouse and
1 s; [* J) M/ J1 o/ T" H( h) ihuman ES cells (Boyer et al., 2005). 2 S- X; P% b4 V8 o) d$ ] t
Given that Klf4 and c-Myc are chro-( _9 ?7 r# X" e& j
matin modifers and can immortal-
! {/ r, B5 U) b0 s% t, j% |( B0 rize cells, one might be able to fnd ; |! x& y4 Q$ {( b4 ?* B0 Z4 @
other factors or small molecules that
! u0 F( C' h9 o( r% Y) T/ |could replace these two factors in the |: M+ \" t8 h) E/ [& @* e
cocktail (Yamanaka, 2007). In these . s8 f# R$ a @
studies, the possibility of retroviral 0 F( L" d u/ h9 K. A
insertional mutagenesis, resulting
' M1 D* e. k# {7 ]3 K: P4 Fin the activation of other genes con-! b% Y3 m; w- v, C @
tributing to reprogramming, cannot
0 ?! D0 C* l5 I+ o0 R- K7 {be excluded, providing an opportu-6 A& v6 V8 M; g" O
nity to potentially identify new repro-; m$ S5 C2 j( _' |: M# |
gramming factors beyond the cur- S6 O' w8 n7 r5 `/ a, V% H1 F
rent quartet. Also, taking a broader . [% c5 i+ a! U* R' H
screening approach for reprogram-
! u4 t% y* W( x7 M; c6 m, Cming human fbroblasts (as Takahashi
( W4 u9 M' k7 a+ ]and Yamanaka did for their mouse
6 \2 X6 m- A; \4 c5 }study) might yield other combinations 8 s- @: U* |$ I( I0 K9 N
of reprogramming factors.
2 }/ }4 L* y4 Y" Z4 m! GDirect reprogramming of somatic
V3 o8 G4 L# j" x: c" }cells to a pluripotent state, thus revers-$ z: F* T" L9 }: ~
ing the developmental arrow of time, 7 s8 A1 ]4 C( {) F3 g; l
is considered by some to be the “holy 9 l1 j! E `# W% h+ h
grail” of stem cell research. Once the
1 l) d9 h3 \5 y: X4 Aresults in human cells are confrmed, & w# v5 n7 L& Z+ {$ }" a9 e
these advances will enable the cre-( I. h$ q$ Q+ I) b, N
ation of patient-specifc stem cell lines
% y& M" x2 s6 J! Qto study different disease mechanisms
; C- X; O2 t+ N+ W9 Hin the laboratory. Such cellular models
( j5 G- B9 I. U# l! t5 nalso have the potential to dramatically $ ]4 Q5 p4 Z' @3 P( O* C4 ~2 H
increase the effciency of drug discov-/ `* V5 @* u* P! k4 w u4 E2 ]
ery and to provide valuable tools for
3 J5 L2 L% L* ^' S1 u8 \toxicology testing. Furthermore, this
" D2 z* b, \/ T3 @, Areprogramming system could make
t$ M5 P7 h4 O2 ?/ ^& L9 jthe idea of customized patient-specifc & y) }* V2 T' a$ p3 b
screening and therapy both possible
, P6 G% J1 ~) s! cand economically feasible. Finally, the * P6 N8 p5 b* P( S
work will have a powerful impact on 8 S' Z2 j' U; e" I) E4 W0 [
the intense debate regarding the moral, * ]& ~: G7 C: `- d! E
religious, and political aspects of ES cell
" ^' y. n0 s! D$ c" Gresearch. However, a big mistake now
! V; l, r- j* [0 O" Xwould be to consider human ES cells , m% }5 w+ P% V7 }
obsolete. There are still many hurdles 5 T" ~, K# X- W
to overcome before we ful ly understand 5 A- I5 u U7 f- ^2 k
pluripotency and before we have human
0 [6 G1 s: `4 [/ G( H( G+ yiPS cells in hand that are suitable for
9 a5 n& t+ [) p2 r- K" t/ Gtherapeutic application. For example, ) A! a; A T ^4 L
a signifcant proportion of mice derived
8 f- T0 A/ G, }: `. _! i7 Ufrom mouse iPS cells develop tumors
, }0 u3 @/ v. C9 ]' ]: Sdue to reactivation of the c-Myc retro-7 s4 H$ M/ Z# w5 D
virus (Okita et al., 2007) compared to
6 n4 Z3 Q/ b- w& i0 [# h# { Ymice derived from ES cells, which are
5 i7 O: P8 Y8 N: wnormal. The search is now on to fnd a % _! a b9 o# L, m; l8 m
way to reprogram somatic cells without
3 y; N" r2 N7 l3 x) V0 s4 bretroviruses and maybe even using a
* ^0 L9 _) ~+ F5 W9 j8 s3 O/ Wcocktail of small molecules. Given this, + N J5 _7 g1 Y' k
it should be emphasized that human
+ ~5 W, b+ \) V5 w8 ?ES cell research is more important than
6 m% W* E/ ^! b3 W1 z$ `ever for it will shed light on how iPS
$ Z# y9 r, J2 u; Q7 ^* F, ncells can best be maintained in their
1 }; t8 k2 R* F& Q, t5 qpluripotent state and how they can be
! r" j( s y) T( Ginduced to differentiate into the cell
5 {# ] j9 _1 }$ P% m# v3 Plineage of interest. The feld of nuclear
! P% D/ m$ @% F6 }7 Q- areprogramming has come a long way
& j0 t* O0 v7 b0 }# Z2 qfrom the initial nuclear transplantation
! _$ o) V8 d! C8 |studies in frogs 50 years ago, to the / ~* [- n# {, ]1 E( f% O( ` Z- q$ h
birth of Dolly, the frst mammal cloned
6 V2 i" x3 z \. v) G5 \from adult somatic cells (Wilmut et al., j" r2 ~2 Q ^
1997), to the fallout from the fabricated & {% z B9 j/ U1 ?# J1 r0 h
human nuclear transfer experiments
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发表于 2009-4-28 21:35
发表于 2009-5-16 10:41
