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Induction of Pluripotency:
* k7 y9 Y# D' i3 d# `8 D, SFrom Mouse to Human
, R" N7 u, P2 f: M1 HHolm Zaehres1( m6 }) p1 o- g1 T
and Hans R. Schöler1,1 `- j6 m9 ]5 l
*
h7 b% U- c7 a4 J8 v( v1
4 o/ }- s: U, \Max Planck Institute for Molecular Biomedicine, Department of Cell and Developmental Biology, Münster, NRW 48149, Germany
: _' l$ k% Z& u*Correspondence: schoeler@mpi-muenster.mpg.de
+ f* V% @ O& L. pDOI 10.1016/j.cell.2007.11.020
: N/ Y; M) h8 R4 K. M. ?; \In this issue of Cell, Takahashi et al. (2007) transfer their seminal work on somatic cell
1 y3 F' Z) z6 breprogramming from the mouse to human. By overexpressing the transcription factor - I5 n( p6 b/ P9 K
quartet of Oct4, Sox2, Klf4, and c-Myc in adult human fbroblasts, they successfully
! p1 J: T/ q/ O$ \% X4 Q( |isolate human pluripotent stem cells that resemble human embryonic stem cells by all " h, L) t& H) w
measured criteria. This is a signifcant turning point in nuclear reprogramming research ( c% u7 T) f4 E: n
with broad implications for generating patient-specifc pluripotent stem cells for research
2 `$ `( ?7 A- Vand therapeutic applications.
i: b6 z @" f' }/ IThis year’s three Physiology or Medi-
: p% ~1 ^2 M( ^; lcine Nobel Laureates—Martin Evans,
* b( j; y+ J( `6 u$ n- vMario Capecchi, and Oliver Smithies—
% h* |% t0 @( i6 \ m9 ]) qwill be honored in Stockholm in 10
2 l3 b, ?9 {: m1 U3 qdays time for their discovery of DNA
5 k/ y4 U: V2 j6 L0 y, |! y* p" ~) _recombination and the development ) w7 u7 b- `% K' X& h% x2 p
of mouse embryonic stem (ES) cell
, @8 q! s* z8 I+ v5 u1 Y5 E0 Ttechnology. It was Martin Evans who # Y* G0 H( O& T, t. ?( t
discovered how to make mouse ES
- i) |. q9 h$ D+ m0 zcells, enabling any genetic alteration F/ |2 r- l2 |( Q$ y/ p7 F
to be transferred to the germline and # b* C: s% z; z* o7 ?; O: x9 I
hence to the next generation (Evans + w, o3 Q, ~$ g- G, P, a+ a
and Kaufman, 1981; Martin, 1981).
1 B: f5 T3 t% `$ J- |Before this breakthrough, researchers 1 u8 o2 e2 e# C& x, s
studied mouse embryonal carcinoma
% h9 }1 Q6 ~- Ecells derived from tumors, which
* `( N1 J: z- O. Wcould form every mouse cell lineage
5 f5 t7 @: h" G. q. lexcept the germline. Combining DNA . Z7 D9 V$ ]. y
recombination and mouse ES cell
0 ]2 ~* `% N- stechnology revolutionized an entire
2 H7 ]- U0 F/ Q$ \4 @feld of research, forming the basis for
" o4 E5 I0 C# d5 o7 X5 k. Sstudying and understanding the roles , ~, S# _7 W/ w8 w
of numerous genes in embryonic . v: d. I3 _6 h7 p$ Z. c r d
development, adult physiology, dis-2 r% ^. o; }7 M' S; C/ ?
ease, and aging. To date, more than
2 p1 }% |/ T* o6 |- _4 `500 mouse models of human disor-9 } V/ ~% T s4 p- |
ders have been generated. Now, with
" {: Y2 B* R9 othe study by Takahashi et al. (2007)
, X( ?' ~% H% U& E0 E& }/ k: Q' ypublished in this issue of Cell, another
4 M# Y# q1 q/ C$ m/ z* dimportant revolution is taking place.
) R3 O- [% e, W% S. LLast summer, Takahashi and
' r8 u/ e$ Y6 I! r7 a4 }& Y" HYamanaka (2006) stunned the scientifc ( E" ]' C& c+ R! g( G# v/ \
community with their study showing
1 m3 L% x8 q% i/ Xmolecular reprogramming of mouse
& m+ K5 e$ a1 isomatic cells into induced pluripotent
% I5 z- R8 m) Hstem (iPS) cells using just four factors:
' I8 V1 [) |2 F r1 S$ OOct4, Sox2, Klf4, and c-Myc. Their 5 x- J- t# p& M ]: D2 H3 y& W
elegant but demanding approach of
0 l6 K2 c1 O+ i2 m- ^% }/ G Fscreening for a cocktail of factors that
r* J4 ?; ?5 M# m7 a! u; A, hcould reprogram mouse fbroblasts
6 l, u# w! q3 [% Fstarting from 24 candidate genes paid % {1 w- x$ q4 G4 y5 A
off with their detailed description of iPS 6 ]5 O3 m7 A9 x" c8 y7 J
cells, which are almost indistinguish-# d4 ^9 z8 F0 Z. e
able from mouse ES cells. As with all
3 _4 |" K4 [! l* ?2 e5 oscientifc discoveries, these exciting
# v% C# _# `- `, O7 v* lfndings had to be reproduced. Sev-
& I! T: C( G7 `, Beral studies published this year not ; v! l! I& O' C
only reproduced but also extended ( g! t* O' f# s
the Takahashi and Yamanaka fndings % a, h( M9 t/ j/ I1 O# g0 C
by demonstrating the pluripotency and ! H4 a, o5 i1 T6 D
differentiation potential of mouse iPS
# J- o; {5 s. x1 bcells in rigorous developmental assays
- B" V$ t" Z; A G u) g1 x, O(Maherali et al., 2007; Okita et al., 2007; 7 K2 j1 r; f6 G- Q, u G7 \1 }
Wernig et al., 2007).
4 ]' Y, e4 i4 iIn their new study, Takahashi, # x+ _1 n$ h2 w- W1 y
Yamanaka, and their colleagues 3 M; k c0 H: Z# ?6 ~
(Takahashi et al., 2007) now translate 7 I7 V) V0 m- v1 |
their remarkable fndings from mouse * f( J" q8 Q# d2 {) u
to human (see Figure 1). They selected # b- O% j" J6 J9 S9 q, |
adult human dermal fbroblasts and ! E, v. ~' g$ ]# \( [& Z
two other human fbroblast popula-
t; P3 f! R1 G0 n4 f1 g% Vtions (from synovial tissue and neo-8 {7 i0 _) z! Y! A" B# Z
natal foreskin) from different human
; {9 D. s' d; r# jdonors as their reprogramming target
9 @) u2 u/ r0 n. }cell populations. They then trans-3 U# u* w9 ]; r9 J' q7 ~# e; W
duced the human fbroblast cultures
. J% A9 i! I9 p/ ]1 K5 b& Wwith retroviral vectors carrying trans-
1 I b1 C3 a$ |9 S3 Y' F, Zgenes for the human versions of Oct4, 6 c% v3 u6 s/ z
Sox2, Klf4, and c-Myc and cultured
2 }" L. d3 ~$ a1 x4 X2 z, qthe cells under human ES cell culture
8 E5 H/ b& _* {* yconditions. Thirty days after transduc-; x, D. P( A6 P
tion, the culture plates were covered
( C! j' q( M! A7 ~: B2 b3 b9 q/ d Nwith human ES cell-like iPS colonies , `# q$ c, o/ z: @
(among other colonies), which could 7 T- G% N+ f. R0 K. G* y9 k7 ~
be further propagated and expanded. : A" Y' w+ v- Z' @0 M
The retroviral vectors enabled silenc-
) [! K; {$ |% b9 Ling of all four transgenes after human . a0 |. s0 R- t) W6 U2 q
iPS formation (as found in the mouse
/ Q4 Q- \4 R6 \0 `; |, F" A( rsystem) indicating that the iPS cells
T- m6 B# y4 w6 [" Dare fully reprogrammed and no longer : f/ C9 \/ T! r: z
depend on transgene expression.$ v2 r, Z5 N1 w
Unlike the mouse study, human - h8 A( B0 L6 {. }, X
iPS cells were generated without any
* Q- F( Y7 s7 |5 Pgenetic selection procedures. Given ! l. C m2 j7 J+ F( W% P
the lower mitotic index of human ES
0 `2 K3 F2 Z0 lcells, it is not surprising that the gen-6 ^2 }9 J Q/ N2 f. W
eration of human iPS cells takes nota-. c" M- W. r& c0 }
bly longer than in the mouse system.
3 \% }( W0 U' ~7 |" ]! zThe authors subjected their human % e3 Z* D1 |) e+ O* u
iPS cells to a panel of assays to com-
' v6 I% y6 X! T- F# ?* ^4 I7 {pare them with human ES cells. These
4 f, w* T3 B) n# p4 \assays included morphological stud-
1 U& j! o1 R, Gies, surface-marker expression, epi-7 N9 v! c+ T) j4 z& ^. j" l: X1 f
genetic status, formation of embryoid
0 T8 Q- H+ U. Y4 Y, Wbodies in vitro, directed differentia-
' \7 ~5 b# T( ^: Ution into neural cells and beating car- I8 J8 R9 G3 b
diomyocytes (according to human
0 S/ |! E; y2 P' a H" D) L1 ~6 }ES cell differentiation protocols), and . T' o+ b7 H. N k% t
fnally teratoma formation in vivo. * R* [$ j5 ^( `% w$ y
DNA microarray analysis revealed 4 s, U" }- ^8 Z( U; P0 ~
the remarkable degree of similar-
9 Y7 b3 H8 j/ Jity between the global gene expres-6 i( K/ B0 g" K3 H9 _1 t
sion patterns of human iPS cells and ?2 F" {5 B: ~8 s {+ e1 M
human ES cells. Notably, genomic
3 t. d% Y# d- }. ZDNA analysis as well as analysis of - ]" O( C Z* \, ?& ?3 `
short tandem repeats demonstrated 1 @" t. h4 d9 V# x0 d
the genetic origin of independent ]" ^" v& T+ {: n" T0 v5 C* d
human iPS clones from their parental " F: y# `) _* J6 W8 A$ U2 S
fbroblast populations.( P- e0 ?, ^+ E, Q
The derivation of mouse and then
( q1 w+ n8 A6 f. {5 O2 s$ thuman ES cells (Thomson et al., 1998) $ q; D) c% ^) T
as the gold standard of pluripotent
" Y& S. c9 T* w2 t: M p7 n8 ?stem cell populations has necessarily
! T& s8 r E# wled to emphasis on differences in the
) {- [* j/ X1 r1 q5 F; Hregulation of self-renewal between , ~& Z# Q& Q! e* h
mouse and human ES cells. For
. ^6 J$ g% u2 J* {( A7 D' dexample, human ES cells depend on
8 i6 m' a$ M$ ybFGF for self-renewal, whereas their * u9 g) e- r* Y
mouse counterparts depend on the # g* N5 b: {$ q2 a; B" s
Lif/Stat3 pathway; BMP is involved in
+ [! {7 v, X) h7 H! smouse ES cell self-renewal, whereas - w9 N: ]1 s4 K/ w7 S
in human ES cells it induces differen-
$ ]2 i) I/ T; i; l ttiation. Extrinsic factors and signals . v- a ^6 E6 K* {& l% R! w
for maintaining pluripotency may dif-+ M4 \) A7 X' g, u5 ^9 s
fer between mouse and human. How-/ T2 S+ g* M: j& j0 \
ever, the ability to translate somatic 4 s: @, k! f: M# M* f6 n1 ^
cell reprogramming from mouse to 4 c3 W; z; o& S4 }- o& |' u# H
human using the same transcription : {+ N. ~2 N1 B( X; C$ A
factor quartet further emphasizes the ! E; V6 w: W+ _& [7 y
conserved nature of the Oct4/Sox2 ; f; F, K1 O' I
transcription factor network that
6 @9 ^) u$ {$ o8 ~& \2 k* zcontrols self-renewal of mouse and
$ B9 a9 d/ l n; ~$ O0 ~human ES cells (Boyer et al., 2005).
+ L9 T* e! V& I( |5 yGiven that Klf4 and c-Myc are chro-1 u* u; h; V) q
matin modifers and can immortal-8 c2 Q) j; l0 |/ K3 z$ x7 J$ m
ize cells, one might be able to fnd 2 b; C* F8 N6 g0 X& G
other factors or small molecules that , `9 x( {+ ?; ^* v
could replace these two factors in the * P/ o! _; b4 g9 I9 B
cocktail (Yamanaka, 2007). In these 8 q$ C; Z# }# \5 ]3 J& ^
studies, the possibility of retroviral
7 i; _8 j4 ~: `9 Z- u' Kinsertional mutagenesis, resulting
; M2 k* l0 M n7 D9 Oin the activation of other genes con-
0 Q/ [1 c) `; J; \7 i' gtributing to reprogramming, cannot
1 S! g* C$ H$ O7 P$ q/ sbe excluded, providing an opportu-* }8 m$ a+ r7 n8 g" N, X: c
nity to potentially identify new repro-
p% b. R9 \' X2 xgramming factors beyond the cur-% ^9 P5 o/ T( l. ^5 d# T
rent quartet. Also, taking a broader
2 L' u- i+ W _+ U# ?9 w5 sscreening approach for reprogram-
& }- i+ ^9 z' ^: {3 x* L0 s: Jming human fbroblasts (as Takahashi 9 C& O* h- [: M2 \0 l8 A( |
and Yamanaka did for their mouse 5 u0 ?' F8 S. r2 N& H; ~
study) might yield other combinations : o% g; N5 X, T: Y8 X) ~
of reprogramming factors.
; G$ X9 R4 k5 A, r: EDirect reprogramming of somatic
1 q* s6 j5 e2 M# T: F- V8 Icells to a pluripotent state, thus revers-% B" [# r( t6 f0 [
ing the developmental arrow of time,
7 j- H5 i2 H7 c8 b) \* b5 o) Ris considered by some to be the “holy
1 E& w+ a# v: Ygrail” of stem cell research. Once the * ]( T9 I0 Q9 S- t9 O. q- D
results in human cells are confrmed, ; e$ o4 [8 m, p0 Q
these advances will enable the cre-
6 |. y3 ]" {2 t0 S& J4 H: @ation of patient-specifc stem cell lines 5 _' b. d S3 J5 M3 k' ]! ]( u
to study different disease mechanisms
! ~0 e ? ~0 r% v& p1 Fin the laboratory. Such cellular models & A5 @7 ^9 y8 _8 `- Y' r+ o. {& Z0 h
also have the potential to dramatically 4 a0 Z- d, \5 l& p# F- C* i
increase the effciency of drug discov-
' n, r+ q5 o: P7 w7 u% V% rery and to provide valuable tools for . v( ~6 R7 |! q. G2 ?
toxicology testing. Furthermore, this
. A2 v5 K! Q! w. ?# Xreprogramming system could make
+ ^# Y+ ]$ M: L1 a9 h. ~0 D0 athe idea of customized patient-specifc : X& o9 C& W0 q2 K* Y$ o3 u4 m4 b
screening and therapy both possible # Y$ ~+ [8 z$ I
and economically feasible. Finally, the
* a p+ W. d, B+ b: v7 V& fwork will have a powerful impact on & L L9 G; }* D& I4 _
the intense debate regarding the moral, 0 n: r s! W; A0 u
religious, and political aspects of ES cell $ @4 T# A y2 ~9 o3 e3 w
research. However, a big mistake now
3 d; y# z& K. e* r9 P' i: Y! Ewould be to consider human ES cells
& {$ g/ n( ]; h- l, z9 |) ]# k* o+ eobsolete. There are still many hurdles
) H* F( ^) _, x1 K5 }0 ~: ?$ fto overcome before we ful ly understand
* K J) Q l: S/ \! w1 E% v) y* D# [- |pluripotency and before we have human
* Q: ]: a6 y5 j6 p; s2 JiPS cells in hand that are suitable for 1 w3 Y7 {# {; G7 C) P
therapeutic application. For example,
8 e9 b4 X$ {, J$ k6 u! ja signifcant proportion of mice derived : R9 A0 U( o. ~! s7 k* t# u
from mouse iPS cells develop tumors
5 h3 f; D+ e, R* t( g# zdue to reactivation of the c-Myc retro-
- @0 ~% J; t. N+ \( vvirus (Okita et al., 2007) compared to 9 n6 z1 ?0 f6 ^4 j9 J
mice derived from ES cells, which are 5 [' q- R( } L$ K4 v6 e5 ?
normal. The search is now on to fnd a - V! \/ Q9 g) F7 b) Q
way to reprogram somatic cells without " u1 g8 M; `4 d" T
retroviruses and maybe even using a ; Q" j3 ]9 X S6 ]# c: @) M
cocktail of small molecules. Given this, $ D! y) Q$ n0 i; n% A0 T
it should be emphasized that human % X: L4 _& S. q& S
ES cell research is more important than
2 q [/ j; F4 X1 c* w; E; never for it will shed light on how iPS
6 ]7 s( m* p& mcells can best be maintained in their
* c# y# B9 S! Q! o& mpluripotent state and how they can be
# d, d) J$ l( G% c5 pinduced to differentiate into the cell
2 u: _2 {, g/ S: Elineage of interest. The feld of nuclear $ r/ N7 a- C! c" X7 r
reprogramming has come a long way " D: z. T' D6 R6 ~; L
from the initial nuclear transplantation ( |! @, c x/ ], V
studies in frogs 50 years ago, to the ( K( m: K) Z3 Y% s* Z
birth of Dolly, the frst mammal cloned 1 o5 b$ V# M9 b3 @% Z/ n
from adult somatic cells (Wilmut et al.,
' W- r( j, k$ C6 g G1997), to the fallout from the fabricated ' n4 H s# B6 {# f" O, B
human nuclear transfer experiments
* Z( X5 d8 o o( X% L. d2 W4 ~ Kof several years ago, to the landmark * F4 z* \& {# k* Z
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their colleagues, frst in mice and now 5 L) i1 @" ]) y9 }. E# R2 b) G
in humans.% Z* J/ S. v. x4 l3 f( A6 \ M8 l
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发表于 2009-4-28 21:35
发表于 2009-5-16 10:41
