Mammary Epithelial Reconstitution With Gene-Modified Stem Cells

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. \2 {% h' m7 E2 g0 l# yThe mammary gland represents a unique model$ |& b5 ?7 D+ r! w
system to study gene functions in adult stem cells.: T% F% \! x2 H8 o2 A7 L: O0 q" l
Mammary stem cells (MaSCs) can regenerate a5 E' k# g6 b. R/ p' v$ n( `
functional epithelium upon transplantation into- l8 \* P+ \. m8 k6 E0 S
cleared fat pads. We studied the consequences of" G+ B' f9 y; K: Z+ C! |
distinct genetic modifications of MaSCs on their6 f. m: y/ N5 v' Z
repopulation and differentiation ability. The
5 `* N. w3 y* a: lreconstitution of ductal trees was used as a stem cell
, @4 v  c# f( h& d8 Rselection procedure and the nearly quantitative, I) k' J  M. y2 `
lentiviral infection efficiency of the primary
2 |( D; }; Q& U+ Omammary epithelial cells (MECs) rendered the
) Y3 Y; t7 |- |enrichment of MaSCs prior to their transplantation
/ G, W8 F5 K- L& ?unnecessary. The repopulation frequency of
$ S' Y5 U+ R4 c- s# Ztransduced MaSCs was nearly 100% in
( N0 V0 ^  }# rimmunodeficient recipients and the resulting7 T( f0 H: k9 C! q) @2 C* r) `
transgenic ducts homogeneously expressed the virally& ~6 s: w5 r- [9 G
encoded fluorescent marker proteins.
* C3 X4 S* {3 QTransplantation of a mixture of MECs, expressing  N; j) a6 e5 u- g$ x6 ]
different fluorescent proteins, resulted in a distinct9 I  q( Q0 p" f" ?5 m
pattern of ductal outgrowths originating from a small7 o1 N7 U; f# t  t5 e3 F( L
number of individually transduced MaSCs. We used
6 _( |. _9 C7 \% G; _* Ygenetically modified MECs to define multiple
; U( Y+ [$ w, q7 H+ ?) f2 Y: Rfunctions of Stat5 during mammary gland
" O4 z7 d4 }0 r; X2 K# P8 ?8 Fdevelopment and differentiation. Stat5-& H* f0 c- a( F5 \, Z4 g" O+ e' O% f
downregulation in MaSCs did not affect primary
, i" g! N9 N/ M4 ]ductal outgrowth, but impaired side-branching and
) o( a2 L2 [9 D; b9 C& s: jthe emergence of mature alveolar cells from luminal0 x- \; g. Q0 z
progenitors during pregnancy. Conversely, the
: |* R& N: d) I( |expression of a constitutively active variant of Stat5,
' z; w- [4 O- o4 q8 E- acS5-F, caused epithelial hyperproliferation,) I* U; Q1 b- R3 s6 o
thickening of the ducts and precocious, functional
4 q/ w0 W' Z4 |& i$ u' K/ E( D) oalveoli formation in virgin mice. Expression of cS5-F
) s, F; D; ]% ?  S" _+ palso prevented involution and caused the formation
4 S0 b: S9 j  {4 t! K5 V( sof ER+PR+ adenocarcinomas. The tumors expressed
! o+ |/ T2 @: \* factivated Stat5 and Stat3 and contained a small/ a+ R( g9 e9 `1 D7 w+ y, p
fraction of CD44+ cells, possibly indicative of cancer: `0 Z" V, s# H( t' [, f; Y$ _: j
stem cells (CSCs)