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Induction of Pluripotency: - g2 r T1 a4 a6 ?, Y! v- g
From Mouse to Human
3 J6 W* \, [6 t( e1 {Holm Zaehres11 B6 x4 t- b& ?+ Q S3 q0 W
and Hans R. Schöler1,
* E3 W+ g, M5 U1 Y" s! {*2 k4 M! h( n7 e7 V0 j1 F
1) i" Y. ]2 B) a% Y
Max Planck Institute for Molecular Biomedicine, Department of Cell and Developmental Biology, Münster, NRW 48149, Germany
2 G1 c6 m8 f- O* w& C3 a*Correspondence: schoeler@mpi-muenster.mpg.de! {" j; A7 e0 ?% y5 J4 a
DOI 10.1016/j.cell.2007.11.0209 `1 e0 Z) p1 a8 Y- a+ d3 u- Q; F; |
In this issue of Cell, Takahashi et al. (2007) transfer their seminal work on somatic cell ' I9 |! X& U a$ j3 d; J1 v
reprogramming from the mouse to human. By overexpressing the transcription factor , V. S0 X; v3 k" K( N: }
quartet of Oct4, Sox2, Klf4, and c-Myc in adult human fbroblasts, they successfully
$ k4 H, \/ E2 j% H" zisolate human pluripotent stem cells that resemble human embryonic stem cells by all % r. k, @9 M4 a# A% D- i$ B
measured criteria. This is a signifcant turning point in nuclear reprogramming research $ v9 q9 u$ H) @0 B; Z6 x+ u6 Y
with broad implications for generating patient-specifc pluripotent stem cells for research
, ~: r/ _6 z- a3 L( c3 H' J* qand therapeutic applications.
! n) g. [4 w. y& H* k. u5 k+ }( B- tThis year’s three Physiology or Medi-
" V/ P+ s; ]/ Ncine Nobel Laureates—Martin Evans, % n n! x1 e7 Q
Mario Capecchi, and Oliver Smithies—
) C$ T: X- S1 a1 V3 z. Iwill be honored in Stockholm in 10
7 {! t: X2 O" p7 S' m* g8 h- i- hdays time for their discovery of DNA
* q* e) _ O( Q wrecombination and the development
2 g1 N6 ]: x0 G1 ~3 o7 Oof mouse embryonic stem (ES) cell % e, c. j' a: p; _! U
technology. It was Martin Evans who - n7 F' t; F# G, u' o
discovered how to make mouse ES
. h& D% T& c; Ncells, enabling any genetic alteration
% G0 p) d3 p+ N5 A E! b- \( Fto be transferred to the germline and * E$ o7 f0 r5 w5 E {: }. {
hence to the next generation (Evans ! E# m- B; o% ^ M( I9 w
and Kaufman, 1981; Martin, 1981).
$ \' [: c3 j4 U( M5 C: k0 EBefore this breakthrough, researchers 6 D3 X- v0 n$ p( `' ~. V
studied mouse embryonal carcinoma
& j$ A# h8 E0 Acells derived from tumors, which 3 @; |7 G; [ s- p3 {+ t
could form every mouse cell lineage
B, p. b* {" Iexcept the germline. Combining DNA
( G: G& H: |( W# o) l3 Srecombination and mouse ES cell ) n! g# ^% w" o+ z
technology revolutionized an entire + V7 P. g1 {3 I0 K
feld of research, forming the basis for
7 d6 @8 G' b* e5 ?; [; Lstudying and understanding the roles
6 v: ]$ @# K1 \5 Y7 y1 Dof numerous genes in embryonic
* @2 D! w n# xdevelopment, adult physiology, dis-
" }# F! r* t/ N; Uease, and aging. To date, more than 6 l+ t7 _* d' n7 c+ F8 v0 B6 p
500 mouse models of human disor-
! w# h3 _; o3 v4 k9 [ders have been generated. Now, with ; w1 f; S0 g3 l Q% _! T; L
the study by Takahashi et al. (2007)
) F3 b9 ?7 g- _published in this issue of Cell, another + [" ?0 m6 k. a' ]3 m
important revolution is taking place. P. g7 m& `9 L. F* S6 P! Q# f( y
Last summer, Takahashi and [- s4 o0 {+ l A) p! j
Yamanaka (2006) stunned the scientifc
8 T* I' q' l3 }( [community with their study showing X0 A$ A1 U/ W% ?' E r
molecular reprogramming of mouse
3 A9 v, r" q2 ~somatic cells into induced pluripotent ! g" X5 n4 M# O$ X+ U2 Q
stem (iPS) cells using just four factors: 9 s( T4 C0 x8 A; X: D {: D
Oct4, Sox2, Klf4, and c-Myc. Their
6 y5 V+ ?9 }) O; w# U: Aelegant but demanding approach of 3 c! R. x' ^7 K
screening for a cocktail of factors that
" C9 I1 a- [* g( T0 o$ |0 Hcould reprogram mouse fbroblasts
: { K6 V( d: t/ f! vstarting from 24 candidate genes paid
- ~& }: g8 _. Y3 Toff with their detailed description of iPS
6 F' S9 r! I8 [( b2 t% ccells, which are almost indistinguish-
* F: i2 q7 c/ Eable from mouse ES cells. As with all ( j2 @; Z6 K2 F- z6 @9 b
scientifc discoveries, these exciting 6 e# }- P. c4 l% c. N7 z
fndings had to be reproduced. Sev-' k% O, S/ F8 Z& f
eral studies published this year not
. \* f N* {4 K$ n0 ]3 w( I3 qonly reproduced but also extended , h5 e8 V/ \3 l% Q8 v& O
the Takahashi and Yamanaka fndings
8 V5 K; a' s# S2 ~" B( @# Kby demonstrating the pluripotency and / @& W4 J1 R. g9 Y& d
differentiation potential of mouse iPS , y3 O/ d5 C7 z$ b- \* }* J& Q
cells in rigorous developmental assays ~% O' z6 O* ~ U, j
(Maherali et al., 2007; Okita et al., 2007;
& E: x/ O' {. \) j7 p$ x! QWernig et al., 2007).
( T3 Q1 Z$ p! N/ r$ UIn their new study, Takahashi,
# l4 I* A7 x4 i9 w5 `% I5 ?0 F, K5 iYamanaka, and their colleagues 8 S& E1 ^! |: k/ r' C) f
(Takahashi et al., 2007) now translate
" U! k$ U5 h: [6 N/ `their remarkable fndings from mouse , }3 p0 C# {6 {) z& I% q' h
to human (see Figure 1). They selected
' g5 ?2 `* l% S- B0 l6 ladult human dermal fbroblasts and
& a7 i% F& |- e% `' y( F) vtwo other human fbroblast popula-
4 O: ?; C- j" R7 dtions (from synovial tissue and neo-! V7 K1 |9 d* Q8 q! ^. y, c
natal foreskin) from different human
5 u4 j9 f- q" j5 O0 l. O$ m. [donors as their reprogramming target : m0 g4 z$ P8 U' p; H5 e- f1 d
cell populations. They then trans-
& \: B8 z4 O6 P! v$ i$ b2 `duced the human fbroblast cultures
" L( Q( B2 `4 ^; k. Owith retroviral vectors carrying trans-* q% [5 _2 z8 M. k4 v4 F, b9 A# c2 O
genes for the human versions of Oct4,
7 n0 O# L* L6 P) zSox2, Klf4, and c-Myc and cultured & V9 |5 U7 v% D
the cells under human ES cell culture : ?, a! q, N) r9 n% j% k$ x
conditions. Thirty days after transduc-* d+ R- E1 _4 ?
tion, the culture plates were covered : E! T2 z2 @* F; B# S) |
with human ES cell-like iPS colonies : `4 X0 `4 P' h' B
(among other colonies), which could 9 \) X# \+ q* }2 ~1 ]- t
be further propagated and expanded.
) N3 Z+ @5 [8 SThe retroviral vectors enabled silenc-9 V( k2 m4 d2 \/ I, d& B9 O
ing of all four transgenes after human S3 j& Z3 |/ g# E8 e: d( e
iPS formation (as found in the mouse
. g1 K+ L T+ P" Z$ j. Tsystem) indicating that the iPS cells ! s' y _1 ~4 a6 Y' F1 U- k- K
are fully reprogrammed and no longer e( K- x8 I2 s& J/ _+ L, I
depend on transgene expression.) J3 l# Q( h: D
Unlike the mouse study, human
. }& m& P* p; R! W: J, \# ziPS cells were generated without any 6 g( U. L; ~4 I1 s- \9 `
genetic selection procedures. Given
! L: p# g& h8 a2 T, i) @# I8 ^the lower mitotic index of human ES - i, a9 V" v: Y
cells, it is not surprising that the gen-3 N* B8 @+ ]4 h/ q
eration of human iPS cells takes nota-
8 |2 R" T8 W! f, n$ u' p9 tbly longer than in the mouse system.
) T6 a; p3 K, [. pThe authors subjected their human
! b) W. M$ N( q& m+ F+ XiPS cells to a panel of assays to com-- K/ @1 T& v/ \" d# {+ r9 ^, O
pare them with human ES cells. These * [, L M9 X8 i4 O
assays included morphological stud-( d! G F, V4 Z1 {( m
ies, surface-marker expression, epi-+ h0 `8 `5 d4 d* ~8 F2 B/ b1 A
genetic status, formation of embryoid : S$ T+ p, n$ ~& `) X- e2 x+ x
bodies in vitro, directed differentia-
- E' g: V9 ~, }* S5 r8 `6 ~tion into neural cells and beating car-
( t4 G4 k0 M p2 p G0 d5 Pdiomyocytes (according to human
% @, t/ m' V8 B. n) d: P: A( e- ^& LES cell differentiation protocols), and
. p' y& k+ C2 ~* _' `fnally teratoma formation in vivo. . N3 v! `) l g; D5 I' q7 ^: {
DNA microarray analysis revealed 8 k6 a# k) v) F
the remarkable degree of similar-3 Z% K/ s7 \0 x+ V9 U/ ^
ity between the global gene expres-
/ a" @" _/ b* I1 G3 t/ {* tsion patterns of human iPS cells and
# k, |+ |5 Y. Z0 x/ _. }human ES cells. Notably, genomic + B5 u7 u3 U2 ^
DNA analysis as well as analysis of ' ?* ~ O* k. M! v! h
short tandem repeats demonstrated
8 f& X! m d: N8 Q2 |+ n& pthe genetic origin of independent
W6 t0 D% U- Vhuman iPS clones from their parental
! c, |' F% i* N% ~4 b$ o. D. m- z8 rfbroblast populations.+ m* Q. A& `3 E* Y# R; s
The derivation of mouse and then
: L- E, {/ d4 S' J' K8 Hhuman ES cells (Thomson et al., 1998) ( w, v9 T" s0 ]3 X0 r7 Y& O2 e
as the gold standard of pluripotent
5 U) J+ K3 k0 F' `' C4 d) Qstem cell populations has necessarily
z: S6 q! J( uled to emphasis on differences in the
& H+ J( C' |2 r' Aregulation of self-renewal between
. {3 M; g5 C2 H# smouse and human ES cells. For
& R; S9 S6 @2 [* i ^& K- q, t6 h! T$ Eexample, human ES cells depend on
! P7 K4 y1 H; W) Z/ a5 K/ rbFGF for self-renewal, whereas their ) q( J( I& A4 C1 N
mouse counterparts depend on the
$ u/ k8 ~! B8 |Lif/Stat3 pathway; BMP is involved in
8 T& W n2 N$ ~% R* J8 {& X# n8 @mouse ES cell self-renewal, whereas $ m0 d" M; G# V3 K7 P8 t
in human ES cells it induces differen-7 n- l8 F n2 n7 Y, I" g
tiation. Extrinsic factors and signals
. @- M8 T9 [$ K7 n' O8 ]for maintaining pluripotency may dif-
! R0 s" g& q. a/ y, ofer between mouse and human. How-+ i8 n" \/ ]. @; K
ever, the ability to translate somatic ! ~ k: y% \; @' U/ P' l
cell reprogramming from mouse to
: j8 V& z4 S# xhuman using the same transcription ' F/ n& E" l: K
factor quartet further emphasizes the
' l' k7 Z6 @ K6 b% v3 nconserved nature of the Oct4/Sox2 . C/ L( `4 q" Y$ d4 P! \
transcription factor network that
& y7 S" s, R- d: h( S, i# rcontrols self-renewal of mouse and ( b m$ e. C# I# {% s" |
human ES cells (Boyer et al., 2005).
- p' l. J5 ^5 M/ X# QGiven that Klf4 and c-Myc are chro-
5 ]. ]4 D; x$ c2 h) C9 Xmatin modifers and can immortal-
+ C5 \* k; s# p1 l8 ] P' Kize cells, one might be able to fnd
1 v" `7 ~& A" Q4 z4 M4 S5 p! p% J0 hother factors or small molecules that 6 P5 y0 _2 ^# Z) E! w! j: d
could replace these two factors in the # p- P! H4 r4 o! @; r) q/ S1 I8 m1 L
cocktail (Yamanaka, 2007). In these ' O; X2 @) B: j2 U9 x$ b4 T" T6 {
studies, the possibility of retroviral
! t' x+ B7 h6 C3 |+ E$ ainsertional mutagenesis, resulting , Q5 o, F; R' R7 L& S
in the activation of other genes con-1 l( o* a( o( l& l( U) N; m) S& S% Q
tributing to reprogramming, cannot & p/ v; y- T. o+ O1 [) o
be excluded, providing an opportu-
& C- l4 P5 b Hnity to potentially identify new repro-
! \9 g! j0 m% kgramming factors beyond the cur-
, X9 l& f9 _ e4 e' Nrent quartet. Also, taking a broader
: t, ^! y8 z, I" Q% J! zscreening approach for reprogram-6 @2 {# o9 U& _0 {# g4 \
ming human fbroblasts (as Takahashi ! ^' W" e: O1 o3 z
and Yamanaka did for their mouse
! d v& B, K ?1 N% `study) might yield other combinations
7 e( G. `( F) @1 u1 t ^$ Q* xof reprogramming factors.
& g$ s/ B& @; C0 O1 u6 I9 z# mDirect reprogramming of somatic
: m, R& K: N* Z3 a1 Gcells to a pluripotent state, thus revers-2 D, u1 ~+ h2 k; Y" Y& w7 D$ y
ing the developmental arrow of time, 8 u: b% p4 J$ c" v
is considered by some to be the “holy
9 ` c$ L5 u" X) Ggrail” of stem cell research. Once the
$ h6 w( m. ?+ f Wresults in human cells are confrmed, 3 }- |9 m, M/ P/ B( l% h. c
these advances will enable the cre-+ v; S+ G4 H( o {* c' D3 y. Q
ation of patient-specifc stem cell lines
% A" h& j, [, L( I& ~to study different disease mechanisms
7 L- D! ~( R. W2 A z1 Vin the laboratory. Such cellular models
6 e" V7 Z4 R7 h, ]8 [- Halso have the potential to dramatically 3 k4 j+ x' m; a7 B& D x" B5 i
increase the effciency of drug discov-* }7 j" w$ q. l% i
ery and to provide valuable tools for
" e7 ^" l) w- J0 U' Dtoxicology testing. Furthermore, this $ I3 o; m; {' G0 Y
reprogramming system could make
?4 h: D5 y, D: _9 Jthe idea of customized patient-specifc
) ?+ X, n/ F) w I" w4 J3 ]screening and therapy both possible
' ]. L# e( n, o* g4 {. c6 gand economically feasible. Finally, the Q3 ^) V! c. N, G" y, h
work will have a powerful impact on / k) \- r' |6 [4 |8 e; k8 p
the intense debate regarding the moral,
4 |% |6 v" u+ n) `6 P' }religious, and political aspects of ES cell
: C5 \7 [- K+ U: x1 ?research. However, a big mistake now
, {4 y# V+ Q3 j- jwould be to consider human ES cells Z( ]* x5 P5 z( ?$ l. Y
obsolete. There are still many hurdles
, D6 T* E! y4 a7 x6 g0 S" K. Kto overcome before we ful ly understand ' x+ ]$ I" o- c; {# Q! m
pluripotency and before we have human : k% S) K3 ]6 }3 R
iPS cells in hand that are suitable for
1 W& B9 b0 @, q' E" Utherapeutic application. For example, 6 c- T @7 U7 z5 Z/ @, t
a signifcant proportion of mice derived 7 g9 y4 W# r2 z
from mouse iPS cells develop tumors ( P6 R8 ?# D x! U
due to reactivation of the c-Myc retro-
2 N/ N/ ^3 o9 O9 w( Qvirus (Okita et al., 2007) compared to 6 ^, f% G$ c% l/ n
mice derived from ES cells, which are
$ q: h! m+ D# r/ X6 m& \! Z( ^normal. The search is now on to fnd a ) v5 p8 _' [2 s. B$ ^
way to reprogram somatic cells without - [& p" D7 C: I
retroviruses and maybe even using a + l% W9 `7 [( s" z* I9 r8 _
cocktail of small molecules. Given this, ( j0 w0 ?# }, S2 e
it should be emphasized that human 8 e7 ^1 D1 k9 J' b0 D% ]/ q, B& I
ES cell research is more important than - N) C0 r, H) ?" f
ever for it will shed light on how iPS
: |# N" J$ w. a0 r- ^cells can best be maintained in their $ q* G( P" \+ s0 L* F* t# A
pluripotent state and how they can be 5 Z% |! C) ?9 y6 Z! A
induced to differentiate into the cell
; g+ e+ ]% \6 p& V7 q4 |lineage of interest. The feld of nuclear 7 F2 h4 ]3 w6 }- W
reprogramming has come a long way
- U1 F7 ^; X6 E O5 p- _+ u% R9 efrom the initial nuclear transplantation : D" c; ]+ b4 U' }1 n
studies in frogs 50 years ago, to the 9 U1 n1 s. |, u; M" R" J
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1997), to the fallout from the fabricated
/ j) y9 ^% |! u- F" ^3 J/ a, M& D5 |human nuclear transfer experiments
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发表于 2009-4-28 21:35
发表于 2009-5-16 10:41
