Formation of a regeneration blastema following limb amputation is believed to occur through a process of dedifferentiation. It has been suggested, however, that the cells contributed to the blastema by the stump muscle are satellite-like cells, rather than cells originated by dedifferentiation. We have previously shown that simple epithelial keratins 8 and 18 are expressed in the mesenchymal progenitor cells of the regenerating amphibian limb and in cultured cells with myogenic potential, and that their expression appears to be causally related to changes in proliferation and differentiation. We show here that retinoic acid (RA) affects the expression of these keratins differently in myogenic cells originated from normal limb and limb blastema. Furthermore, we find that the effects of RA on proliferation, myogenic differentiation and adhesion of these cells also differ. In fact, whereas RA does not affect keratin expression, proliferation or myogenic differentiation in blastemal cells, it does decrease keratin levels and thymidine incorporation and increase myogenesis in cells from normal limb. Conversely, RA increases cell adhesion only in blastemal cells. Significantly, these effects of RA on cultured cells are consistent with those observed in vivo. Overall the results presented here suggest that in the urodele limb there are two distinct cell populations with myogenic potential, one originating from dedifferentiation and one equivalent to the satellite cells of the mammalian muscle, which are likely to be primarily involved in blastema formation and muscle repair, respectively.

REFERENCES

Alric
S.
,
Froeschle
A.
,
Piquemal
D.
,
Carnac
G.
,
Bonnieu
A.
(
1998
).
Functional specificity of the two retinoic acid receptor RAR and RXR families in myogenesis.
Oncogene
16
,
273
282
Brockes
J. P.
(
1997
).
Amphibian limb regeneration: rebuilding a complex structure.
Science
276
,
81
87
Brown
R.
,
Brockes
J. P.
(
1991
).
Identification and expression of a regeneration-specific homeobox gene in the newt limb blastema.
Development
111
,
489
496
Cameron
J. A.
,
Hilgers
A. R.
,
Hinterberger
T. J.
(
1986
).
Evidence that reserve cells are a source of regenerated adult newt muscle in vitro.
Nature
321
,
607
610
Carlson
B. M.
(
1970
).
Relationship between the tissue and epimorphic regeneration of muscles.
Am. Zool
10
,
175
186
Casimir
C. M.
,
Gates
P. B.
,
Ross-Macdonald
P. B.
,
Jackson
J. F.
,
Patient
R. K.
,
Brockes
J. P.
(
1988
).
Structure and expression of a newt cardio-skeletal myosin gene. Implications for the C value paradox.
J. Mol. Biol
202
,
287
296
Corcoran
J.
,
Ferretti
P.
(
1997
).
Keratin 8 and 18 expression in mesenchymal progenitor cells of regenerating limbs is associated with proliferation and differentiation.
Dev. Dyn
210
,
355
370
Epstein
L. M.
,
Gall
J. G.
(
1987
).
Self-cleaving transcripts of satellite DNA from the newt.
Cell
48
,
535
543
Ferretti
P.
,
Brockes
J. P.
(
1988
).
Culture of newt cells from different tissues and their expression of a regeneration-associated antigen.
J. Exp. Zool
247
,
77
91
Ferretti
P.
,
Brockes
J. P.
(
1990
).
The monoclonal antibody 22/18 recognizes a conformational change in an intermediate filament of the newt, Notophthalmus viridescens, during limb regeneration.
Cell Tissue Res
259
,
483
493
Ferretti
P.
,
Brockes
J. P.
,
Brown
R.
(
1991
).
A newt type II keratin restricted to normal and regenerating limbs and tails is responsive to retinoic acid.
Development
111
,
497
507
Ferretti
P.
,
Fekete
D. M.
,
Patterson
M.
,
Lane
E. B.
(
1989
).
Transient expression of simple epithelial keratins by mesenchymal cells of regenerating newt limb.
Dev. Biol
133
,
415
424
Ferretti
P.
,
Geraudie
J.
(
1995
).
Retinoic acid-induced cell death in the wound epidermis of regenerating zebrafish fins.
Dev. Dynam
202
,
271
283
Fuchs
E.
,
Green
H.
(
1981
).
Regulation of terminal differentiation of cultured human keratinocytes by vitamin A.
Cell
25
,
617
625
Geraudie
J.
,
Ferretti
P.
(
1998
).
Gene expression during amphibian limb regeneration.
Int. Rev. Cytol
180
,
1
50
Gordon
H.
,
Brockes
J. P.
(
1988
).
Appearance and regulation of an antigen associated with limb regeneration in Notophthalmus viridescens.
J. Exp. Zool
247
,
232
243
Griffin
K. J. P.
,
Fekete
D. M.
,
Carlson
B. M.
(
1987
).
A monoclonal antibody stains myogenic cells in regenerating newt muscle.
Development
101
,
267
277
Halevy
O.
,
Lerman
O.
(
1993
).
Retinoic acid induces adult muscle cell differentiation mediated by the retinoic acid receptor-alpha.
J. Cell Physiol
154
,
566
572
Hay
E. D.
(
1959
).
Electron microscopic observations of muscle dedifferentiation in regenerating Amblystoma limbs.
Dev. Biol
1
,
555
585
Jing
Y.
,
Zhang
J.
,
Waxman
S.
,
Mira-y-Lopez
R.
(
1996
).
Upregulation of cytokeratins 8 and 18 in human breast cancer T47D cells is retinoid-specific and retinoic acid receptor-dependent.
Differentiation
60
,
109
117
Kim
K. H.
,
Stellmach
V.
,
Javors
J.
,
Fuchs
E.
(
1987
).
Regulation of human mesothelial cell differentiation: opposing roles of retinoids and epidermal growth factor in the expression of intermediate filament proteins.
J. Cell Biol
105
,
3039
3051
Kintner
C. R.
,
Brockes
J. P.
(
1984
).
Monoclonal antibodies identify blastemal cells derived from dedifferentiating muscle in newt limb regeneration.
Nature
308
,
67
69
Lane
E. B.
,
Bartek
J.
,
Purkis
P. E.
,
Leigh
I.
(
1985
).
Keratin antigens in differentiating skin.
Ann. NY Acad. Sci
445
,
241
258
Lo
D. C.
,
Allen
F.
,
Brockes
J. P.
(
1993
).
Reversal of muscle differentiation during urodele limb regeneration.
Proc. Natl. Acad. Sci. USA
90
,
7230
7234
Maden
M.
(
1983
).
The effect of vitamin A on the regenerating axolotl limb.
J. Embryol. Exp. Morph
77
,
273
295
Navarro
J. M.
,
Casatorres
J.
,
Jorcano
J. L.
(
1995
).
Elementscontrolling the expression and induction of the skin hyperproliferation-associated keratin K6.
J. Biol. Chem
270
,
21362
21367
Ohtsuki
M.
,
Tomic-Canic
M.
,
Freedberg
I. M.
,
Blumenberg
M.
(
1992
).
Regulation of epidermal keratin expression by retinoic acid and thyroid hormone.
J. Dermatol
19
,
774
780
Oshima
R. G.
,
Abrams
L.
,
Kulesh
D.
(
1990
).
Activation of an intron enhancer within the keratin 18 gene by expression of c-fos and c-jun in undifferentiated F9 embryonal carcinoma cells.
Genes Dev
4
,
835
848
Oshima
R. G.
,
Baribault
H.
,
Caulin
C.
(
1996
).
Oncogenic regulation and function of keratins 8 and 18.
Cancer Metast. Rev.
15
,
445
471
Powell
A. J.
,
Gates
P. B.
,
Wylie
D.
,
Velloso
C. P.
,
Brockes
J. P.
,
Jat
P. S.
(
1998
).
Immortalization of rat embryo fibroblasts by a 3-untranslated region.
Exp. Cell Res
240
,
252
262
Scadding
S. R.
(
1989
).
Histological effects vitamin A on limb regeneration in the larval axolotl, Ambystoma mexicanum.
Can. J. Zool
68
,
159
167
Schrag
J. A.
,
Cameron
J. A.
(
1983
).
Regeneration of adult newt skeletal muscle tissue in vitro.
J. Embryol. Exp. Morph
77
,
255
271
Semat
A.
,
Vasseur
M.
,
Maillet
L.
,
Brulet
P.
,
Darmon
Y. M.
(
1988
).
Sequence analysis of murine cytokeratin endo A (no. 8) cDNA. Evidence for mRNA species initiated upstream of the normal 5end in PCC4 cells.
Differentiation
37
,
40
46
Tanaka
E. M.
,
Gann
A. A. F.
,
Gates
P. B.
,
Brockes
J. P.
(
1997
).
Newt myotubes reenter the cell cycle by phosphorylation of the retinoblastoma protein.
J. Cell Biol
136
,
155
165
Thoms
S. D.
,
Stocum
D. L.
(
1984
).
Retinoic acid-induced pattern duplications in regenerating urodele limbs.
Dev. Biol
103
,
319
328
Thornton
C. S.
(
1968
).
Amphibian limb regeneration.
Adv. Morphogen
7
,
205
249
This content is only available via PDF.