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Induction of Pluripotency: ! ~4 s8 _* y0 `( M
From Mouse to Human% ]2 A, J' g. [
Holm Zaehres1- H/ ?! a& U; ?* }3 i1 x
and Hans R. Schöler1,- J8 R( z2 e$ c, _9 D5 m- H* }
*
. q+ Y9 |' C9 g4 Z' I# Z* ^$ }3 N1
Q3 z( R+ e& V( GMax Planck Institute for Molecular Biomedicine, Department of Cell and Developmental Biology, Münster, NRW 48149, Germany
6 b4 L) i3 v7 \9 W' q7 ~*Correspondence: schoeler@mpi-muenster.mpg.de
: q6 f% t x$ n2 f) y% `5 r6 }DOI 10.1016/j.cell.2007.11.020
9 d/ o- q5 r7 \' ~1 i! e6 _( KIn this issue of Cell, Takahashi et al. (2007) transfer their seminal work on somatic cell
3 X$ X8 e/ h9 `reprogramming from the mouse to human. By overexpressing the transcription factor
( G1 V9 O* x0 Y) d: a1 I3 P+ jquartet of Oct4, Sox2, Klf4, and c-Myc in adult human fbroblasts, they successfully
8 B) A z6 V% @+ f9 e7 T. y/ {! aisolate human pluripotent stem cells that resemble human embryonic stem cells by all
6 }7 E: {) B3 s9 jmeasured criteria. This is a signifcant turning point in nuclear reprogramming research
2 E' [6 A6 B, q. Bwith broad implications for generating patient-specifc pluripotent stem cells for research
9 T. }6 X8 [5 f$ @" d2 F7 \and therapeutic applications., W4 _& ]$ m5 w; c8 v/ Z' ?1 r, f
This year’s three Physiology or Medi-8 q8 z- G; Z6 G9 d6 R
cine Nobel Laureates—Martin Evans, % n* C. K1 p% N7 I1 \- e' n
Mario Capecchi, and Oliver Smithies—
& s( l" X+ q1 y0 w; s& \will be honored in Stockholm in 10 ! a4 Y# N7 \3 v. ^! o- l7 |' \
days time for their discovery of DNA % ]/ N) p3 e$ c- A8 u7 ]
recombination and the development
! m# d2 `. z0 F1 L2 R, Bof mouse embryonic stem (ES) cell
8 k* F: n7 B5 U' ftechnology. It was Martin Evans who
1 I1 j0 n; G" }5 ^% Q. rdiscovered how to make mouse ES
- R% v. O i# B7 Z- T6 \- Xcells, enabling any genetic alteration
% B& L; T6 Q2 B7 P' o$ ?to be transferred to the germline and
- Y1 A) Q4 F# d# r2 ^7 ghence to the next generation (Evans
: o1 _/ @0 _0 |% w+ f/ J; `and Kaufman, 1981; Martin, 1981).
- c2 I4 k# [- {Before this breakthrough, researchers
4 G8 l! M* a6 \) G. }+ G: y( h7 @+ istudied mouse embryonal carcinoma
6 U5 \6 G$ G. h1 p k5 y$ Hcells derived from tumors, which $ M0 X _2 ~9 Q$ r; n$ B
could form every mouse cell lineage # h( O# l6 i+ M) \" V
except the germline. Combining DNA 0 Y# H: X' T! s# q7 G
recombination and mouse ES cell
9 z8 e O. ^ l4 | a) etechnology revolutionized an entire # b. X1 {. i/ Z& J# F
feld of research, forming the basis for ' v6 n& F6 p, B
studying and understanding the roles - y* ?3 h6 z1 F6 `+ B1 f0 F
of numerous genes in embryonic
, l- v: p8 d1 G8 Jdevelopment, adult physiology, dis-. A+ c9 ^! D; }
ease, and aging. To date, more than + d+ X. }* c' k! K0 K
500 mouse models of human disor-* `- \# i/ Z% H: z/ p$ o
ders have been generated. Now, with / W6 ^3 c) @7 L% T! t' Q! c/ b7 X
the study by Takahashi et al. (2007)
; F- k. Y4 Q' |" H6 Q, l9 {9 n: ipublished in this issue of Cell, another 9 n1 ~( H0 v# Q2 ?* ~/ u4 ?
important revolution is taking place. \) B! K- ^1 x) b, W! u
Last summer, Takahashi and
# ~5 m6 q6 B7 Y3 @5 DYamanaka (2006) stunned the scientifc
- |* p1 h+ a D* ?# Rcommunity with their study showing
, n) @) D% ]- R$ Smolecular reprogramming of mouse 8 l. G' z2 k. i2 u: h: ^) T' } x
somatic cells into induced pluripotent
, i; C' W( a! b6 }1 P% h8 Wstem (iPS) cells using just four factors:
0 ~$ ]+ h$ Q+ LOct4, Sox2, Klf4, and c-Myc. Their
2 h* A6 b6 ] @% O# belegant but demanding approach of
. D$ A$ U. F! q( n9 Escreening for a cocktail of factors that 8 {7 R$ _; k. O
could reprogram mouse fbroblasts ; m' P, F W9 g$ n1 i
starting from 24 candidate genes paid 1 n. s! y' z5 }0 @
off with their detailed description of iPS 6 _8 @! j' \" e$ G
cells, which are almost indistinguish-! w) A N! u( F1 f, r- ~ l* N
able from mouse ES cells. As with all ' s. M6 ^# F' h! t2 x J
scientifc discoveries, these exciting ' z. v/ m; `& G% m1 g. g
fndings had to be reproduced. Sev-# x0 F" x- M( B" n) O& D. X
eral studies published this year not - \ ]- q, \- X3 x
only reproduced but also extended ) c' X. s9 m' z' e2 E- D
the Takahashi and Yamanaka fndings
' l2 A# d( g9 t- @4 hby demonstrating the pluripotency and z4 f; p. m% ]9 S- w q
differentiation potential of mouse iPS $ A6 }* S. z f0 ` n! o9 A. ]
cells in rigorous developmental assays , T. T) R2 c5 i1 A7 I0 Q$ e3 p0 o
(Maherali et al., 2007; Okita et al., 2007; 7 p' |4 c+ I9 ^
Wernig et al., 2007).
1 [! k( ~- C/ l+ B9 P* I4 A2 X# _In their new study, Takahashi,
3 @6 `1 X4 E6 f- r! A% m% `Yamanaka, and their colleagues
R6 h& C# Q w$ P; p& _(Takahashi et al., 2007) now translate
- q* q/ |/ B- a F; ^8 Ptheir remarkable fndings from mouse ; k- U) G7 E& D9 a# ]
to human (see Figure 1). They selected
4 W) L s. m. f6 xadult human dermal fbroblasts and + p$ X& e4 Q2 y. ~3 A
two other human fbroblast popula-
1 c m% s# T- Y: M& D* Utions (from synovial tissue and neo-- F. L* J s0 B5 T$ {( n
natal foreskin) from different human ' I9 X' b( U- R; y+ P0 m1 c
donors as their reprogramming target 1 e7 b& X7 k* ?, T
cell populations. They then trans-7 Z9 ]1 L' G+ K/ z
duced the human fbroblast cultures $ \# _$ }# T _0 G H0 _0 t
with retroviral vectors carrying trans-% d) }( V" v1 h) d
genes for the human versions of Oct4, 2 u8 g3 k S1 w- B6 V( @
Sox2, Klf4, and c-Myc and cultured : a) t5 o7 P' k! t
the cells under human ES cell culture
! j. S/ L( Z8 t# u6 P& oconditions. Thirty days after transduc-
) r) j0 p/ p- C5 Otion, the culture plates were covered 5 k, k" }5 K! w0 M0 ^4 b
with human ES cell-like iPS colonies $ z0 T4 Y2 r0 B, S- a: j2 l
(among other colonies), which could
" p w5 u2 v' x; h5 T1 C6 Fbe further propagated and expanded. % D" ^* L! v' T7 U5 u
The retroviral vectors enabled silenc-
4 u& a0 |0 K% m( Ying of all four transgenes after human n+ }+ ~% E6 r3 x
iPS formation (as found in the mouse ! Y9 H: [- H: g! C# i$ c
system) indicating that the iPS cells `+ W T% D$ I/ S2 K- U; x% p1 S
are fully reprogrammed and no longer
4 [, g0 { k- [: I& Qdepend on transgene expression.( W: l/ u+ w+ N
Unlike the mouse study, human & W+ Y2 ]4 v, P! R
iPS cells were generated without any 0 o: g. G2 H# T4 I9 }
genetic selection procedures. Given " F1 S5 ~0 o0 m" x
the lower mitotic index of human ES
8 g3 Z2 F- z' G: }2 i9 Dcells, it is not surprising that the gen-% x+ F! h! C2 w' o4 q. g5 O( e
eration of human iPS cells takes nota-
) T: |% Q7 V# Z0 N- Nbly longer than in the mouse system.
8 f& Q; R) a x% D/ { k; N8 |The authors subjected their human
' e$ q8 v2 q G) U5 ~iPS cells to a panel of assays to com-
L/ y2 c6 v- xpare them with human ES cells. These + b* q, L4 y6 H/ n0 m3 \
assays included morphological stud-6 s. S+ s6 \. ^7 O. F& E
ies, surface-marker expression, epi-
$ G* N r9 q$ g W U2 }9 T' w' p9 W& ogenetic status, formation of embryoid
7 o' S1 ~8 f2 L3 ?3 d% |bodies in vitro, directed differentia-
: `7 Y9 a. W w8 z8 r+ @6 L6 jtion into neural cells and beating car-* W$ K7 h7 w; K# |: d9 ]7 X: E" m
diomyocytes (according to human " ^' w# L, U }' T9 W
ES cell differentiation protocols), and
/ |7 `( r/ R% a% mfnally teratoma formation in vivo.
7 }/ T* L% P7 w& G/ ^9 IDNA microarray analysis revealed
, a9 _+ a% x5 s: B& ~' E# ~: sthe remarkable degree of similar-
0 ^6 N2 t, f# ^/ l( y0 S! Qity between the global gene expres-
) X# f0 c. p3 K; I w3 {sion patterns of human iPS cells and
0 M' K% n3 v! X; Q2 t! N9 Bhuman ES cells. Notably, genomic # z' f! K3 [" z0 I5 K
DNA analysis as well as analysis of . c, n/ ~) k; L
short tandem repeats demonstrated % O; `$ W2 f+ r& ?! G' S
the genetic origin of independent
4 j! j/ Z+ v% g, e5 X* m! Dhuman iPS clones from their parental , {% [) |$ i( _; ]/ {0 R. B
fbroblast populations.; B5 j7 v& _$ P$ d; l, ]) P% Y
The derivation of mouse and then / [, H9 m% a/ z: ?1 r- o O$ F$ P
human ES cells (Thomson et al., 1998) + F; n0 V( D0 I( l1 g& y4 X
as the gold standard of pluripotent 9 c3 m. \8 M$ j0 U5 t$ }
stem cell populations has necessarily
* D* p0 S4 W9 eled to emphasis on differences in the
. ]( }. [8 u& w( Kregulation of self-renewal between
. j2 N. B" `. Smouse and human ES cells. For 7 ]# S5 y6 x3 E+ m8 ?! @( y! {
example, human ES cells depend on ; b. c3 h4 F& C2 a3 l i
bFGF for self-renewal, whereas their
- C7 V( v/ J2 w- qmouse counterparts depend on the " y& j) O0 j/ Q* O2 {
Lif/Stat3 pathway; BMP is involved in $ S4 O+ d4 c5 q$ D# q+ j- ?
mouse ES cell self-renewal, whereas
6 ?. p a% M6 `) i. p# I1 `; Xin human ES cells it induces differen-
, E; t) e+ q0 A' B; j' T% m! ftiation. Extrinsic factors and signals
5 c- l0 y1 F' s: c% C( D3 z- Pfor maintaining pluripotency may dif-
, [/ @# {5 r& b8 E6 b+ wfer between mouse and human. How-
8 o- m5 y$ P5 {6 S# V/ ?$ H: ?ever, the ability to translate somatic
* a/ i. \ Z; V; Ocell reprogramming from mouse to ' s$ }0 O3 M/ u- b
human using the same transcription + z# H; u& N9 d$ r
factor quartet further emphasizes the 4 |4 ?& A, c- n. x% K# ~4 [
conserved nature of the Oct4/Sox2
: W- ?4 d7 W* m: N8 Y2 ^% {: ytranscription factor network that 6 J. A% K! |4 U& N
controls self-renewal of mouse and ) Y$ a. |4 w2 ^8 Z! c5 c! l( |1 ^
human ES cells (Boyer et al., 2005). 5 b- y' O3 E7 T; d$ z: p
Given that Klf4 and c-Myc are chro-9 m# K( @" a" Z5 d/ n% `
matin modifers and can immortal-2 n' p* k8 Q- t: e; Q, h# J: F2 f5 o' ?
ize cells, one might be able to fnd
6 k+ i' ]/ q# N# q: m) @other factors or small molecules that 7 o# m9 s! i, ~3 s8 C2 g/ \0 [' p
could replace these two factors in the 8 U- n/ O( t1 Z9 W3 Y$ ]0 P0 j
cocktail (Yamanaka, 2007). In these
4 k3 V* H% @6 z Dstudies, the possibility of retroviral ' J# ? j: s) s# i; Q1 Z* b
insertional mutagenesis, resulting
+ R$ t9 _, I# Y: r2 @# rin the activation of other genes con-+ S1 O: T( ? l9 ]' J" Z% ]; a
tributing to reprogramming, cannot . S* R: ]+ k! m
be excluded, providing an opportu-% C3 K' }- A; A. b. C* B
nity to potentially identify new repro-' R" L2 o5 a' { D; ?
gramming factors beyond the cur-2 }- W# K- U# I- I7 x7 V( t/ X
rent quartet. Also, taking a broader
1 L/ y {% {) C1 R. Escreening approach for reprogram-9 W9 G0 [) c/ i. n% [
ming human fbroblasts (as Takahashi
- {- a1 c5 p2 ?$ |* }and Yamanaka did for their mouse
& Z5 Y) t3 m- Q. ^study) might yield other combinations
$ _% s/ b5 v* ?4 W8 qof reprogramming factors.
5 u* S. b) s: b, v/ q' k# U9 Q( RDirect reprogramming of somatic
& V1 f7 e" D4 U7 J6 [& wcells to a pluripotent state, thus revers-$ s, Z% S5 f: @2 @/ D- I
ing the developmental arrow of time,
! X/ ~7 I9 a# Y4 J; g; {7 |is considered by some to be the “holy
2 l% L9 Q0 T8 bgrail” of stem cell research. Once the
u" ~9 O$ s0 h- d0 ~results in human cells are confrmed,
+ s9 P0 u/ L5 w$ M* Jthese advances will enable the cre-
+ d9 I( V/ b6 p4 `& ration of patient-specifc stem cell lines
3 r$ U4 K9 ^1 Z1 X3 V7 z+ M- Jto study different disease mechanisms
& b8 P& D. g6 S# B" _& h |in the laboratory. Such cellular models
" p) h3 ^, w( }: B3 {also have the potential to dramatically * H8 I9 P) T7 |, a9 a" m4 S
increase the effciency of drug discov-
' E. [5 }% Z7 ^9 ^$ n6 B1 F/ z2 N: uery and to provide valuable tools for
, {) V) ^6 s* K7 Q3 z+ n8 _7 ktoxicology testing. Furthermore, this 0 f& A( G+ ?2 e: M( e6 B
reprogramming system could make ! I& v2 ~6 g0 S* o# p* s
the idea of customized patient-specifc
- J" S" q; H( n. O7 bscreening and therapy both possible
8 H' w6 _3 `# k7 z8 ~and economically feasible. Finally, the 4 ~( u1 A4 t0 P8 A+ I" V5 v
work will have a powerful impact on . l) n# S3 y U% L+ V0 V3 L
the intense debate regarding the moral, + R/ ~7 }+ e$ K
religious, and political aspects of ES cell
" X( L& s" l p: a N2 }1 fresearch. However, a big mistake now
( t) z! E# ^2 u' |would be to consider human ES cells
) j$ o" O% S& w* U) Fobsolete. There are still many hurdles
* x1 o4 f3 o5 u7 \3 W1 Bto overcome before we ful ly understand ! }6 r R1 n2 G/ V% c
pluripotency and before we have human
0 N+ N# z1 X0 h% z3 ZiPS cells in hand that are suitable for + e( O k3 A7 G/ H3 Z/ f1 f" F
therapeutic application. For example,
2 j* ~/ r& N8 j3 O; z& ?a signifcant proportion of mice derived
! M# Q6 ~' z' [. m+ A. P8 }from mouse iPS cells develop tumors * F' ^: p c. _; A
due to reactivation of the c-Myc retro-6 S& Y% A: P* o: B4 T& e
virus (Okita et al., 2007) compared to ' l! y% |! H8 h O! ~9 @7 \( j4 D
mice derived from ES cells, which are / T9 D# _' r1 q9 Y" C
normal. The search is now on to fnd a
/ D7 i _( {; _& U4 d/ ^way to reprogram somatic cells without 7 ^- N+ q8 K% s- g6 S [( L; @) G
retroviruses and maybe even using a : [( T, I* \+ [6 }+ M% \
cocktail of small molecules. Given this, ; t3 @8 L; H3 K5 ?+ e+ v
it should be emphasized that human
4 C6 E( N) a7 S2 _ES cell research is more important than
% {9 t% |' ~8 B& [9 Xever for it will shed light on how iPS
, {: l! k1 R) `+ qcells can best be maintained in their
* ^& ~' `( w8 t( {0 @/ fpluripotent state and how they can be
8 F) C5 v( F: J; C9 ]induced to differentiate into the cell
% \ M+ u6 Q0 H7 G, f- @9 ^# a8 rlineage of interest. The feld of nuclear 2 N: m3 x! N) X2 q3 M
reprogramming has come a long way 3 K# P* P' R8 {0 n6 K: o( {) `7 C
from the initial nuclear transplantation $ X- Q( _5 E: s* j `8 s, M
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0 f" }8 F+ M5 b" Y3 i) k3 [, u7 Jbirth of Dolly, the frst mammal cloned ) n! W" }6 [( [
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发表于 2009-4-28 21:35
发表于 2009-5-16 10:41
