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Induction of Pluripotency:
_" Y7 X" n1 i& d6 i2 z! e2 wFrom Mouse to Human0 i1 H0 j. d2 g1 M1 t% K a
Holm Zaehres1
* k2 q) V6 Y' s% \ and Hans R. Schöler1,
1 F+ @; J* g+ r, Z0 |8 m o*% |) S/ J# Q7 d3 I
1
E$ q: s4 ?. h% E2 g+ @8 RMax Planck Institute for Molecular Biomedicine, Department of Cell and Developmental Biology, Münster, NRW 48149, Germany2 p; w* R% l' Q; o! R
*Correspondence: schoeler@mpi-muenster.mpg.de: V1 U5 ~2 S- H1 k
DOI 10.1016/j.cell.2007.11.020
6 N; |+ A) W. u4 ~0 D* A% l) q0 K/ NIn this issue of Cell, Takahashi et al. (2007) transfer their seminal work on somatic cell 6 c7 f2 P/ O& B
reprogramming from the mouse to human. By overexpressing the transcription factor ; s6 i0 T: K, B; ?! D
quartet of Oct4, Sox2, Klf4, and c-Myc in adult human fbroblasts, they successfully
4 N3 l- y0 F0 C9 I% H8 Zisolate human pluripotent stem cells that resemble human embryonic stem cells by all
8 T$ o$ C+ v- ?% }7 [0 Wmeasured criteria. This is a signifcant turning point in nuclear reprogramming research
) q/ b, m' p+ f$ h" A4 b) |8 gwith broad implications for generating patient-specifc pluripotent stem cells for research 4 L: d6 C# N% ~' G5 C
and therapeutic applications.4 m0 r. b4 e; k) Q' f; @2 W
This year’s three Physiology or Medi-5 v2 C5 J; I# a( ?
cine Nobel Laureates—Martin Evans, ' x S" d3 o. O# I2 L# `( r5 J
Mario Capecchi, and Oliver Smithies—
, M3 _2 G+ O+ N7 _6 c; x5 B2 ^will be honored in Stockholm in 10 H7 e$ L& H: t) Y" o* `
days time for their discovery of DNA " @6 G+ V. Z$ e% E
recombination and the development
$ u: M) ^9 {3 }( ~! D0 @of mouse embryonic stem (ES) cell & H8 l% C: P. e. c. Y4 R0 k
technology. It was Martin Evans who
4 n' g" D! {1 O: T( R/ mdiscovered how to make mouse ES
' }4 n( q; E. k5 ?+ V x9 T( pcells, enabling any genetic alteration
# Z% p- m, |$ ?to be transferred to the germline and
, `+ {! h' a9 p4 o; w9 qhence to the next generation (Evans , B0 c( {0 `6 w0 ~8 n! u
and Kaufman, 1981; Martin, 1981). + r/ Q+ B2 [' [. Q+ V
Before this breakthrough, researchers $ O: r. P( V( f& ]; e
studied mouse embryonal carcinoma
( Y: q3 f6 u, W) W6 ocells derived from tumors, which 3 F z- Y% A- ]7 @
could form every mouse cell lineage
! S& \3 t) W3 }except the germline. Combining DNA
, k: ~# }" F! z, q: A. L7 urecombination and mouse ES cell
& i t5 n7 N- u* [/ m0 g' h6 J7 ^technology revolutionized an entire , K* b2 W4 u! H7 E
feld of research, forming the basis for . z1 J0 _8 L" p6 F5 b5 N
studying and understanding the roles
. m' ~) x( P$ ?. B. oof numerous genes in embryonic 4 y8 i- y2 X: s$ ?( e
development, adult physiology, dis-7 Y% h- e7 |2 n0 _* S- X
ease, and aging. To date, more than
# ]/ Q5 N# Q6 e: L7 Z& P500 mouse models of human disor-5 X/ _/ o* x" J1 q" s- x6 x! Y
ders have been generated. Now, with , |$ y/ }; d; k
the study by Takahashi et al. (2007)
) _/ p- j0 p9 y6 e8 {; @2 U) tpublished in this issue of Cell, another
1 E8 ]2 U5 W+ X0 dimportant revolution is taking place.- `) o% f6 M# i- C, O# g( p
Last summer, Takahashi and % \$ ~9 k6 a' c
Yamanaka (2006) stunned the scientifc # G: t. y. k2 E1 m7 Q' l
community with their study showing & `% Z9 R( E$ D. Z4 ~$ ~! `
molecular reprogramming of mouse
5 {. g, B7 E) r& {3 Esomatic cells into induced pluripotent
, M; n9 D5 b- e7 rstem (iPS) cells using just four factors: : _9 j5 m, R$ j8 v( b; B
Oct4, Sox2, Klf4, and c-Myc. Their 5 n5 |3 O$ `& W3 o
elegant but demanding approach of 9 k. B+ O* P$ H2 W/ h: |3 Z
screening for a cocktail of factors that
/ U. Y X( Y! T. E9 qcould reprogram mouse fbroblasts $ t/ s. U( X( l6 L8 r+ m
starting from 24 candidate genes paid
( c4 `/ ?; X9 g5 I Yoff with their detailed description of iPS : K! g0 k% A& m# `$ c1 u
cells, which are almost indistinguish-/ w; o+ F" T1 s0 f: C1 M6 {; m
able from mouse ES cells. As with all
( E3 O8 T8 }* q9 R/ Y6 t, Cscientifc discoveries, these exciting ' F1 P) F5 U* x6 P1 Y1 M* P* \0 ?
fndings had to be reproduced. Sev-
8 a4 g: b1 d0 T' d. _eral studies published this year not 2 k5 L6 [' Z% n' U. T- S5 j
only reproduced but also extended & s. l8 V8 @' i3 l7 F, C2 i+ H7 }$ l
the Takahashi and Yamanaka fndings 2 H0 `& q" ~0 m6 u9 q
by demonstrating the pluripotency and $ z( U7 ]1 H& U, E; c# ~! E; y
differentiation potential of mouse iPS * x9 i& B% e5 U1 C' n7 ~
cells in rigorous developmental assays
4 b/ b. Z6 @5 V; u(Maherali et al., 2007; Okita et al., 2007; P# b3 H+ F( W; y1 n$ O& N
Wernig et al., 2007).
+ }/ `/ d" ~0 |4 \6 t9 F% XIn their new study, Takahashi, 6 K6 O+ }8 G- l4 }+ ?) W% L
Yamanaka, and their colleagues + p# r+ ?! ^+ n( g l, [) I
(Takahashi et al., 2007) now translate
; r; j8 F8 i! htheir remarkable fndings from mouse 0 _) b9 J6 f1 S; i G
to human (see Figure 1). They selected
2 H; I9 d/ E L; [5 A4 b- l8 kadult human dermal fbroblasts and
1 e. f7 ^ j; Btwo other human fbroblast popula-5 e1 X! S6 |" j( c/ n
tions (from synovial tissue and neo-
^; A/ n+ v" {- c7 dnatal foreskin) from different human [1 d( O7 q# ? N% q+ r- l
donors as their reprogramming target
4 s" J# W/ m) xcell populations. They then trans-4 ?4 R" v8 K; i T7 ?
duced the human fbroblast cultures ) H; L. ]* ], a6 d
with retroviral vectors carrying trans-
E# N) W+ p7 Z, ~/ j& G5 |# Tgenes for the human versions of Oct4, " d+ C9 [5 a0 q: [" m5 j) I
Sox2, Klf4, and c-Myc and cultured
5 H/ ^0 d! i9 zthe cells under human ES cell culture % Q% V: j5 E5 S f2 b3 E
conditions. Thirty days after transduc-3 }( Y" R& k# @( b; } N
tion, the culture plates were covered . h! G/ s; s" Y( [
with human ES cell-like iPS colonies
6 n: Y3 g1 U- O, V(among other colonies), which could
$ N1 u: E8 Z3 x" I3 u* pbe further propagated and expanded.
5 u. b9 R9 u z/ U; d& f: yThe retroviral vectors enabled silenc-
! u C8 o. J1 \5 X6 D3 V6 _ing of all four transgenes after human & d& c9 |$ \; j& J' ?6 h; C0 u# k, Y
iPS formation (as found in the mouse
; d; S/ A4 c7 D6 V( Ssystem) indicating that the iPS cells : ~2 v7 k* {# D4 G3 I# [- @# K" ^
are fully reprogrammed and no longer 8 B& s y+ y: o7 U
depend on transgene expression.
- s# C9 T% X9 n. o" o% SUnlike the mouse study, human
$ y, u, T' N1 }iPS cells were generated without any ! @' ?5 {8 L5 M# \8 Z# t
genetic selection procedures. Given
1 }8 }$ f5 O3 U- G N# pthe lower mitotic index of human ES
9 z) v4 `5 m' v% Fcells, it is not surprising that the gen-
0 D8 {+ g- w8 O: veration of human iPS cells takes nota-( G) X+ Z. |! W( Z5 h4 s
bly longer than in the mouse system.
0 N7 p, s0 ?% pThe authors subjected their human 0 B! O& E3 `4 a0 j5 z% w' i
iPS cells to a panel of assays to com-" Y" f! S- B& g3 J8 Q) k# K1 [' b
pare them with human ES cells. These
/ V# t( \( Z( S' o8 Z( m2 v* F( zassays included morphological stud-: e! s) n9 O+ y6 Y5 U" _
ies, surface-marker expression, epi-) G- g5 }& ]. H# j6 w4 g8 L
genetic status, formation of embryoid
3 i% H/ \# w& U( t) R2 fbodies in vitro, directed differentia-
( a. t9 j$ p( B1 k) t! Qtion into neural cells and beating car-
' |0 r3 V8 a/ cdiomyocytes (according to human
* }6 f' |5 M$ Q. T. W6 X) C* P% ^" `! p. pES cell differentiation protocols), and
( z; T9 \9 M% h. A# Tfnally teratoma formation in vivo. ' ]+ q9 U. E; \ F% i
DNA microarray analysis revealed
; H" a" {$ A& y8 N; N( Gthe remarkable degree of similar-% D' [, F* [" S" z: d" k) T: W
ity between the global gene expres-
. Y7 }4 ^7 p7 x. d' _sion patterns of human iPS cells and r/ x- T7 I% D, z" V9 P0 ~- t
human ES cells. Notably, genomic
" H7 d; n8 j3 u2 ^; I: w' BDNA analysis as well as analysis of . \+ z9 X) u% y# ]" _
short tandem repeats demonstrated
2 Q0 |8 Z! ^; H5 Zthe genetic origin of independent
8 f& m; b9 E0 V! r G( Phuman iPS clones from their parental * h! y% _* H; W" k' l) e
fbroblast populations.
& G* A6 [1 r8 |& v2 ]0 RThe derivation of mouse and then 3 @8 h% E. D* j: i9 T `
human ES cells (Thomson et al., 1998) - @* ^" J$ C% d$ p6 P& C; q
as the gold standard of pluripotent 6 C w! v) w3 r5 } C/ \
stem cell populations has necessarily 3 D! H$ P2 O5 x4 |& _) b8 b
led to emphasis on differences in the 5 k0 R0 D/ _. K
regulation of self-renewal between 5 S U( q: x1 G, x; Z
mouse and human ES cells. For ; m2 Q- T8 F) \# r3 ~
example, human ES cells depend on & A9 c# e3 X' k& p d f
bFGF for self-renewal, whereas their ; m. ?) {' b! F6 ]' h
mouse counterparts depend on the 9 A2 `. A& j" F! x
Lif/Stat3 pathway; BMP is involved in
2 M( O J$ K& n/ c% _/ {3 N/ u: u Cmouse ES cell self-renewal, whereas
# N# w& [6 ~0 b5 i H8 Zin human ES cells it induces differen-. Z! d3 \: \: j+ X4 f+ Z
tiation. Extrinsic factors and signals 0 ~+ I0 q) a/ f# F9 w5 E; n
for maintaining pluripotency may dif-
$ L8 ^) T+ n. Yfer between mouse and human. How-+ { S* E4 \+ ^( x
ever, the ability to translate somatic
: c9 S; Y0 d: b E# }/ ycell reprogramming from mouse to 5 B: Z+ V0 I" ^0 S
human using the same transcription
6 o; p: k8 A! _) Vfactor quartet further emphasizes the
& m7 y) F; O! Gconserved nature of the Oct4/Sox2
& Q3 a1 D; u1 T1 A' o- ^transcription factor network that
. k1 [6 ^% t* P. ~controls self-renewal of mouse and
8 O5 X4 W4 y. f% b4 V7 Fhuman ES cells (Boyer et al., 2005). 4 h% I8 ?3 s- i$ H: b& T
Given that Klf4 and c-Myc are chro-7 g0 q/ U( _2 k1 ?2 {
matin modifers and can immortal-, }! {. V1 x9 b- w; J4 c" m
ize cells, one might be able to fnd
. n, U' d$ B( } O2 y1 K. kother factors or small molecules that # W% J* b. b: z! {
could replace these two factors in the * d$ U% K9 i9 r _; U5 }
cocktail (Yamanaka, 2007). In these 5 k2 H0 C" J/ K# t
studies, the possibility of retroviral 0 T/ X& _. |! t! d( ]
insertional mutagenesis, resulting ! @# Z/ ^5 A* L
in the activation of other genes con-6 l0 R( P& w* }2 p
tributing to reprogramming, cannot
; z# x7 o7 Q% c8 U; Kbe excluded, providing an opportu-, N! y% {2 W5 \- X
nity to potentially identify new repro-& f7 B- I. p/ x: I& L6 E) l+ [
gramming factors beyond the cur-* y) r, J6 c4 B C+ }/ M; ?
rent quartet. Also, taking a broader # L* R/ ]" {4 {+ |. m
screening approach for reprogram-: C/ L$ Q. `: J# r# T
ming human fbroblasts (as Takahashi
2 C$ j- A3 ?, C- `# ]and Yamanaka did for their mouse
1 {" e2 z* Q" k. f# R! b/ L2 U$ Ustudy) might yield other combinations 8 @; `* a* m9 y4 {7 [, `7 q
of reprogramming factors.
' m. k# G, [ {0 }! w7 x% y: iDirect reprogramming of somatic
" I$ R' S7 O0 g8 N' Q' x, J0 Kcells to a pluripotent state, thus revers-# \) q9 r& I8 Z, Y$ {( M, i4 v9 J
ing the developmental arrow of time, : U9 p( T: r) i
is considered by some to be the “holy
* N9 V, J& \! J2 J# m2 _& c6 Igrail” of stem cell research. Once the
2 O* {7 V& l; J& E3 xresults in human cells are confrmed,
8 L& v6 g' M8 X8 L0 t5 sthese advances will enable the cre-0 o2 _1 p. z/ g7 R9 ?. a
ation of patient-specifc stem cell lines
# M, {+ A3 B/ x5 W: Y' Eto study different disease mechanisms
0 \! C& _! c! H+ i" sin the laboratory. Such cellular models & y& v& b0 y' b9 K0 K: m
also have the potential to dramatically " b5 }& m+ A4 J9 k9 V
increase the effciency of drug discov-5 U( T, t; a: L' T9 T, F/ w
ery and to provide valuable tools for % w, ^( e6 f; G* g. M% O1 ^
toxicology testing. Furthermore, this
# ^1 c! q# L' Q6 p8 Q; \reprogramming system could make ; W2 F5 l V# A$ U3 d
the idea of customized patient-specifc
! y$ \) k4 F6 H$ r) A# Ascreening and therapy both possible
) j/ _. @+ O& \& v- dand economically feasible. Finally, the
* X! L% [7 F: w0 \: X# Kwork will have a powerful impact on
2 y4 {4 u- e0 mthe intense debate regarding the moral,
: T0 q6 _$ f( s0 x areligious, and political aspects of ES cell % A0 ]9 u( N! l: ?
research. However, a big mistake now 7 E- `5 z- j8 v8 S5 a5 M
would be to consider human ES cells ! n7 l# \1 z* r0 D% ^$ p: p
obsolete. There are still many hurdles $ L0 O% e9 @9 L9 T, s j. X
to overcome before we ful ly understand
+ V( s) D, f9 S+ _* Upluripotency and before we have human
! `. v& i& x; ?, `" j$ u F2 qiPS cells in hand that are suitable for
% T3 y/ E. {% J+ }5 Z4 t0 Dtherapeutic application. For example, . V( B, ~7 b2 d* k5 R9 D
a signifcant proportion of mice derived 0 U# B* y% U$ \3 x, `
from mouse iPS cells develop tumors . k5 U7 c# C0 D# D8 q$ N
due to reactivation of the c-Myc retro-
8 U( ]6 m+ W Cvirus (Okita et al., 2007) compared to $ @7 S% a& D7 |2 }
mice derived from ES cells, which are
5 ?1 J3 O7 [ O2 g2 N( h; @, tnormal. The search is now on to fnd a
9 u, Z) K$ E9 Gway to reprogram somatic cells without
! z6 Y9 ]/ F5 g7 Z7 P, vretroviruses and maybe even using a
) x7 [2 W- y8 D+ ~: H8 E- Ecocktail of small molecules. Given this,
: e- L0 ?& k! I) a& C7 Bit should be emphasized that human - i( A% C3 a0 J$ C4 ?3 _
ES cell research is more important than # n2 _( W6 m/ H `# f- m/ Y( N4 S
ever for it will shed light on how iPS
2 U2 g$ l- L3 Ycells can best be maintained in their
n9 u5 k& B; s+ S J0 i1 X* mpluripotent state and how they can be ( C' e1 i4 R* ~( ]
induced to differentiate into the cell 4 n) v! ?( i, d9 ~1 R+ S
lineage of interest. The feld of nuclear % C4 }" C( {8 `( {/ s
reprogramming has come a long way 3 x& z/ s1 f# W7 ?
from the initial nuclear transplantation 9 e k% Y& G& u/ ^
studies in frogs 50 years ago, to the
: `: u5 p/ ~4 P& X$ gbirth of Dolly, the frst mammal cloned
* I; J G6 }7 s" Z. @; ifrom adult somatic cells (Wilmut et al., 0 J( q4 _% L% ^# {- E Q9 ^% V
1997), to the fallout from the fabricated " a) a( t/ x3 [. j
human nuclear transfer experiments
; {$ f! a1 G7 ^% ~of several years ago, to the landmark ' X, ?6 t! g) r" V m6 `
studies of Takahashi, Yamanaka, and
% v) `1 M% ]2 \% a. o" x. ~2 `their colleagues, frst in mice and now ! Y# O, O% v2 Z
in humans.; h. ]( y. `. [# G0 r. _) C0 i( n: H
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发表于 2009-4-28 21:35
发表于 2009-5-16 10:41
