Reviews of Reproduction (2000) 5, 189–199

Role of progesterone in peripheral nerve repair

H. L. Koenig* W. H. Gong* and P. Pelissier
Laboratoire de Neurobiologie du Développement, Université Bordeaux 1, 33405 Talence, France

Progesterone is synthesized in the peripheral nervous system in glial cells. The functions of
progesterone are indicated by the findings that it stimulates neurite outgrowth from dorsal
root ganglia sensory neurones in explant cultures, accelerates the maturation of the regen-
erating axons in cryolesioned sciatic nerve, and enhances the remyelination of regenerated
nerve fibres. The formation of myelin sheaths around axons is a sexually dimorphic process, as
the sheaths are thicker in female than in male regenerating nerves. The progesterone-induced
myelination is probably mediated by progesterone receptors, as it is impaired by mifepristone
(RU486), a progesterone antagonist. The stimulation of neurite growth in the peripheral
nervous system may be mediated by a progesterone metabolite, 5α-tetrahydroprogesterone,
through GABAA receptors.

Progesterone is conventionally thought of as a female gonadal
steroid hormone synthesized in the ovary. Its classical target
tissues are the uterus, mammary glands and brain. There is
now abundant evidence that progesterone is more than just
a female sex hormone. It is also a neurosteroid synthesized in
the central and peripheral nervous systems, where its non-
reproductive functions are only beginning to be understood
(Koenig et al., 1995; Schumacher et al., 1996; Baulieu, 1997; Jung-
Testas et al., 1999). This review summarizes the evidence that
progesterone has neurotrophic roles in the peripheral nervous
system (PNS), that it activates the growth and maturation of
axons and stimulates the repair and replacement of myelin
sheaths in regenerating nerve fibres. Progesterone, as has been
suggested for androgens, may be a ‘potential therapeutic agent
in enhancing the reparative response of neurones to injury’
(Jones, 1993).
The possibility that steroid hormones are also involved in
nerve regeneration was first indicated by the finding that
inhibition of the synthesis of cholesterol, the precursor of
steroid hormones, inhibits neurite outgrowth in retinal explants
(Heacock et al., 1984), delays myelination in the sciatic nerve
of postnatal mice (Rawlins and Uzman, 1970) and that the
treatment of crushed rabbit nerves with steroid hormones
accelerates muscle re-innervation (Vita et al., 1983). This review
reports studies of previously unrecognized neurotrophic roles
of progesterone in the peripheral nervous system: (1) in acti-
vating the growth of axons and their maturation, and (2) in
stimulating the repair and replacement of the myelin sheaths
in regenerating nerve fibres.
Progesterone in the peripheral nervous system
Classically, two experimental systems are used in the study of
the synthesis or function of molecules such as progesterone in
*Present address: Laboratoire de Génétique Moléculaire de la
Neurotransmission et des Processus Neurodégénératifs, CNRS UMR
C9923, Bât. CERVI, Hôpital de la Pitié-Salpétrière, 75013 Paris, France.
© 2000 Journals of Reproduction and Fertility
1359-6004/2000
the PNS. First, in vitro, dorsal root (DRG) and sympathetic
ganglia explants or dissociated cells, incubated in the presence of
trophic factors, are used to assay ‘neurite-promoting activity’.
Second, in vivo, the influence of neurotrophins and neuro-
trophic factors, including progesterone, is analysed mainly in
the lesioned peripheral nerves of adult animals After a nerve is
wounded (cut, crushed or locally frozen), the distal part of the
axons degenerate rapidly (Fig. 1).
Simultaneously, the myelin sheaths break down within the
Schwann cells, which are then phagocytosed by the macro-
phages that invade the degenerating nerve. New Schwann cells
migrate to the lesioned zone and multiply rapidly inside the
basal lamina tubes. The basal lamina tubes are maintained after
freezing, as they are after crushing of the nerve but are not main-
tained after its section (Mira, 1979, 1988). In the distal stump,
the unfrozen Schwann cells divide actively while degenerating
axons and myelin sheaths are removed by macrophages. These
events are referred to as ‘Wallerian degeneration’ (Fig. 1). Axonal
sprouts, which result from the branching of the proximal un-
severed axons, appear rapidly in the frozen zone, and are en-
closed in the basal lamina tubes at the external surface of the
degenerating Schwann cells. The axonal sprouts surrounded
by new Schwann cells constitute the ‘regenerating axonal
bundles’. Next, the diameter of one or two sprouts become
larger (approximately 2 µm). These sprouts segregate from the
bundles and form separate ‘single axons’, surrounded by
Schwann cells in the usual 1:1 relationship. During this process,
myelination is initiated, either inside the bundles or in the sepa-
rated ‘single axons’ (Fig. 1). This sequence of events, which
mimics the developmental process, may be called ‘axonal
maturation’ (Scherer and Salzer, 1996).
The first evidence that progesterone is synthesized in the
PNS comes from the finding that pregnenolone, the precursor
of progesterone, is present in large quantities in the sciatic
nerves in rats (Akwa et al., 1993) and humans (Morfin et al.,
1992). In mice, both pregnenolone and progesterone accumu-
late in the peripheral nerve (Koenig et al., 1995).
In vitro, [3H]pregnenolone is converted to [3H]progesterone in
Schwann cells harvested from rat embryonic dorsal root ganglia
190 H. L. Koenig et al.

progesterone (Guennoun et al., 1997). Neurones of DRG in-

Muscle fibre cubated in [3H]pregnenolone produce significant amounts of
Axon Myelin Cryode Basal [3H]progesterone and [3H]5α-dihydroprogesterone. In similar
(a)
lamina conditions, Schwann cells isolated from DRG produce radio-
active progesterone, 5α-dihydroprogesterone and 3α,5α-
tetrahydroprogesterone (Guennoun et al., 1997). Rat sciatic
nerve fragments were shown to contain 5α-reductase, the en-
zyme that converts progesterone to 5α-dihydroprogesterone
Motoneurone Nucleus (Celotti et al., 1992).
body Schwann Node of Motor
cell body Ranvier endplate
Effect of progesterone on axonal growth and maturation in
the peripheral nervous system
(b)
Studies in vitro
The addition of progesterone to cultured DRG from 16-day-
old rat embryos results in the growth of a halo of neurites sur-
Internode Invading SC rounding the explants and increased myelination (Fig. 2).
Macrophage Both the surface area and the apparent density of the
neurites increase (compare Fig. 2a with b). The surface area
of the growing neurite increases by 60–130% after 3 days of
Unmyelinated Axonal progesterone treatment (Fig. 2e). Progesterone may stimulate
SC sprouts neurite growth through progesterone receptors present in
(c)
DRG (Jung-Testas et al., 1999). Low amounts of progesterone
receptor have been detected in rat peripheral nerves (Jung-
Testas et al., 1996) but not in mouse sciatic nerves (I. Jung-
Testas and H. Koenig, unpublished). The addition of RU486
Promyelin Growth to control cultures of DRG containing no progesterone did not
fibre cone decrease the growth of neurites (W. H. Gong and H. Koenig,
unpublished). Therefore, it is uncertain whether the endogen-
ous progesterone produced within DRG plays a role in neurite
(c) Remyelinating SCs outgrowth. The addition of 5α-dihydroprogesterone and 3α,5α-
tetrahydroprogesterone to cultured DRG also increased neurite
outgrowth (W. H. Gong and H. Koenig, unpublished). Since
3α,5α-tetrahydroprogesterone does not bind to nuclear pro-
Reinnervation of gesterone receptors, but interacts with gamma-aminobutyric
muscle fibre acid (GABA)A receptors (Celotti et al., 1992; Melcangi et al.,
1999), it is possible that 3α,5α-tetrahydroprogesterone stimu-
lates neurite outgrowth via this receptor subtype. This view is
supported by the finding that the stimulation of neurite out-
Fig. 1. The processes involved in the regeneration of peripheral growth from DRG explants by 3α,5α-tetrahydroprogesterone,
nerve fibres after damage by local freezing (cryolesion). (a) An un- 5α-dihydroprogesterone and progesterone is inhibited by the
lesioned nerve. The myelinated Schwann cells are separated by GABAA antagonists: bicuculline and picrotoxin (W. H. Gong
nodes of Ranvier, but the basal lamina is continuous up to the
and H. Koenig, unpublished). Furthermore, progesterone-
motor end plate. Schwann cells are responsible for myelin for-
induced neurite outgrowth from DRG explants is blocked by
mation. (b) A few days after the nerve is damaged, the Schwann
cells, myelin sheaths and axons degenerate distally to the site of in- finasteride, an inhibitor of 5α-reductase. Therefore, it appears
jury. Macrophages invade the lesioned area and the distal part of that neurite outgrowth is stimulated by 5α-dihydroprogesterone
the nerve to phagocytose the degenerating debris. (c) New Schwann or 3α,5α-tetrahydroprogesterone, or both, rather than by
cells appear in the cryolesioned zone, and multiply in this zone and progesterone (W. H. Gong and H. Koenig, unpublished).
in the distal stump of the nerve. The proximal unfrozen regenerat- Sensory neurones in DRG are heterogeneous cells with dif-
ing axons end with growth cones and axonal sprouts issued from ferent cell body sizes, functions and receptors. Small cells
these axons, which grow inside the basal lamina tubes. (d) After ap- mediate pain and temperature, express trk A receptors and
proximately 1 week, the regenerated axons are re-ensheathed and are sensitive to nerve growth factor (NGF). Medium-size cells
remyelinated by Schwann cells in the cryolesioned nerve. The target
mediate pressure sensations, express trk B receptors and re-
muscle cells are also reinnervated.
spond to brain-derived neurotrophic factor (BDNF) and neuro-
trophin 4–5. Large cells mediate proprioception, express trk C
and respond to neurotrophin 3. The question arises if all
explants (Koenig et al., 1995). In DRG cultures, both Schwann or only subpopulations of DRG neurones are progesterone-
cells (satellite cells) and neurones contain 3β-hydroxysteroid and 3α,5α-tetrahydroprogesterone-sensitive and repond to
dehydrogenase (3β-HSD), which converts pregnenolone to steroids.
 Role of progesterone in peripheral nerve repair 191

Fig. 2. Effects of progesterone on neurite growth and myelination in explanted dorsal root ganglion
(DRG) sensory neurones. (a–b) Neurite outgrowth in embryonic rat DRG explants taken at day 16 of
embryo development. After a 2 day treatment with cytosine–arabinoside (Ara-c), progesterone was
added to the control culture medium, which permanently contained 10 ng NGF ml–1 to allow survival
and a minimal outgrowth of neurites. In control medium, 24 h after Ara-C treatment, the neurite out-
growth is poor (a); after addition of 100 nmol progesterone l–1 for 24 h, the neurites occupy a significantly
larger surface area which grows further (b). (c–d) Myelination in rat DRG explants taken at day 17 of
embryo development. Myelin was obtained according to the method of Eldridge et al. (1987), modified by
DoThi (1992), as described in Koenig et al. (1995). Myelin was stained with Sudan black. After 4 weeks in
Sato’s defined medium, the ganglia were cultured in a myelin-promoting medium containing ascorbic
acid for 2 more weeks. In the presence of 20 nmol progesterone l–1 (d) the total length and density (mm–2)
of the myelinated fibres were increased approximately sixfold compared with control dishes (c). Several
nodes of Ranvier are visible in the cultures (arrow). G: dorsal root ganglion explant. (e) Kinetic of neurite
outgrowth (expressed in mm2) in control and progesterone-treated (100 nM) DRG explants. Both types
of cultures contained 10 ng NGF ml–1 for 5 days (2 days Ara-C treatment plus 3 days of experiment).
(f) Density of myelin (expressed in µm mm–2) measured 15 days after progesterone (20 nM) was added to
the myelinating culture medium . **P < 0.001 (ANOVA Student’s t test).

mutant Trembler mouse (Koenig et al., 1991). The Trembler has

Studies in vivo
an autosomal dominant mutation of the myelin protein PMP22
Studies in vivo on the effects of progesterone on axonal (Suter et al., 1992). Mutant mice develop tremor, quadriparesis
maturation have been carried out in the sciatic nerve from the and transient seizures during early development (Falconer,
192 H. L. Koenig et al.

Fig. 3. Effect of progesterone on the maturation of regenerating axons in the hypomyelinated peripheral nerve of a Trembler male mutant
mouse. Electron micrographs from a series of ultrathin transverse sections through the sciatic nerve. The sciatic nerves were cryolesioned
by repeated freezing (6–8 cycles of freezing–thawing) with a 2 mm copper cryode previously frozen in liquid nitrogen. Progesterone
(100 mg kg–1), dissolved in sesame oil, was immediately applied onto the nerves. After 2–4 further applications, the nerves were dissected
out and fixed for electron microscopy after 7, 12 and 20 days, respectively. (a) Control sham-operated nerve, 7 days after cryolesion and
application of the vehicle (sesame oil). Most regenerated axons (ax) and the sprouts (s) they give rise to are grouped in dark Schwann cells.
This structure forms the axonal bundles (thick arrows), which are surrounded by multilayered basal laminae, which characterize the
Trembler peripheral nerve fibre (thin arrows). There were no myelinated axons in the nerves examined. (b) Progesterone-treated nerve,
7 days after cryolesion. The number of single axons in a 1:1 relationship with Schwann cells increases as a consequence of the retraction of
the sprouts. Some axons (ma) are already surrounded by a thin myelin sheath (arrowhead). (c) Progesterone-treated nerve, 12 days after
cryolesion. The number of axonal bundles is markedly reduced, whereas myelination is increased. (d–e) Sciatic nerves, 20 days after cryo-
lesion: (d) control (non-treated) nerve; (e) progesterone-treated nerve. The mean thickness of Trembler regenerated myelin sheaths is ap-
proximately 15% lower in control nerves than in progesterone-treated nerves.
 Role of progesterone in peripheral nerve repair 193

(a) (b)
60 4.0
Clusters of regenerated fibres 3.5
50

Axons (number per cluster)

3.0
40 ** ++
2.5

30 2.0
*** 1.5
20
** 1.0
10
0.5

0 0
7 12 7 12
Days after cryolesioning Days after cryolesioning

(c) (d)
30 10
*
***
25 *
8
Mean number of lamellae
Myelinated fibres (%)

++
20
6
15
4
10

2
5

0 0
7 12 20 7 12 20 Unoperated
Days after cryolesioning Days after cryolesioning control

Fig. 4. Effect of progesterone applications to cryolesioned sciatic nerve on axonal regeneration and
myelination in Trembler mutant mice. The sciatic nerve was exposed and a portion subjected to local
freezing. Progesterone and control vehicle (sesame oil) were then applied several times for 7, 12 and
20 days. Measurements were made directly on the electron microscope screen or on the electron micro-
graphs. The Trembler mice used in these experiments were heterozygous adult mice (homozygous mutants
do not survive in our strain). (a) Number of axonal bundles present after 7 and 12 days after cryolesion in
progesterone-treated (■; n = 8) and sesame oil-treated (control; ■; n = 5) regenerating nerves. (b) Number
of axonal sprouts per axon bundle. (c) Number of myelinated fibres. (d) Number of myelin lamellae.
*** P < 0.001; ** P < 0.01; * P < 0.05; ++ P > 0.01 (ANOVA Student’s t test). (a–d) The results were obtained
in the same groups of animals (7, 12 and 20 day old mice.

1951). The mutation produces severe hypomyelination and de-
myelination of the peripheral nerve fibres. The number of
Schwann cells in peripheral nerve fibres is 8–10 times higher
than in normal mice. Each Schwann cell is surrounded by multi-
layered basal laminae (Ayers and Anderson, 1973; Koenig et al.,
1991; Do Thi et al., 1993; Fig. 3a–d). The Trembler mouse has two
features that make it useful for experimentation. First, 20% of the
sciatic nerve axons only are thinly myelinated in cross-sections
of the nerve. These axons degenerate rapidly, within 24–48 h in
the areas subjected to cryolesioning (Koenig et al., 1991). Thus,
the regenerating axonal sprouts and subsequent myelination are
easy to see. The rate of axonal regeneration after cryolesion of
194 H. L. Koenig et al.

Progesterone Progesterone

Schwann cells Schwann cell

nuclear receptor of PR DHP,THP THP 5α-DHP
progesterone 3α,5α-THP
Neurone
Neurotrophic soma
factors Neurone GABAA-R

Signal to axon
(membrane or axoplasm) SC GABAA-R Axon GABAA-R

Retrograde signal to Hyper polarization

neurone soma (axon membrane)
Gene activity
Signal to axon
Retrograde signal to
Neuronal
Axonal growth neurone soma
gene
Gene activity
activity
Axonal maturation
Axonal growth Axonal maturation

Myelin initiation

Myelin wrapping

Fig. 5. Hypothetical mechanisms explaining the direct stimulatory effect of progesterone on myelination in the Schwann cells of regen-
erating peripheral nerve fibres. PR: progesterone receptor; DHP: dihydroprogesterone; THP: tetrahydroprogesterone; GABAA-R: GABAA
receptor.

the sciatic nerve in Trembler nerves is double that in normal mice
(Ferzaz et al., 1989). Second, cryolesioned axons in Trembler
mice have robust regenerative capabilities (Ferzaz et al., 1989).
Axonal sprouts first appear 6–12 h after cryolesion and remain
enclosed in the Schwann cell sheath, surrounded by the basal
lamina tube, for more than 12 days. Local application of exogen-
ous progesterone to the cryolesioned sciatic nerve of the Trembler
mouse accelerates the process of regeneration. The number of
clusters of regenerating fibres (Fig. 4a) and the number of axons
per cluster decreases in the bundles of treated nerves (Fig. 4b).
Twelve days after cryolesion, the axonal bundles and the
number of their sprouts are still lower in progesterone-treated
nerves than in the untreated control nerves (Fig. 4a,b). As a
consequence of the loss of redundant sprouts and bundles, the
number of individualized ‘single axons’ doubles in 7 days in
progesterone-treated nerves (Fig. 3c).
It is not known whether the progesterone produced in
the Trembler sciatic nerve is involved in axonal growth and
maturation, since the concentrations of progesterone and its
precursor, pregnenolone, are very low (Koenig et al., 1995).
Axonal maturation promoted by exogenous progesterone is
probably the result of an acceleration of the natural re-
generative processes. It is not known whether 5α-dihydro-
progesterone and 3α,5α-tetrahydroprogesterone accelerate
axonal maturation in vivo. The mechanism by which axonal
growth is stimulated by progesterone or its 5α-derivatives
probably involves several neuronal genes expressed in neuro-
filaments, microtubules and in the axolemma. The expression
of these genes may be regulated either at transcription or at
translation level by progesterone or its 5α-metabolites. The
progestagen target cells may be neurones or Schwann cells.
In regenerating nerves, Schwann cells synthesize and secrete
several neurotrophic factors that contribute to axonal growth
(Terenghi, 1999). Progestagens may enhance the synthesis of
neurotrophins or other factors that induce an axonal signal for
growth. The signal molecule may act either locally on the
axonal membrane or the growth cones or be transported in a
retrograde direction to the neurone cell bodies to stimulate the
genes of proteins associated with and necessary to the growth
of the axons (Fig. 5).
 Role of progesterone in peripheral nerve repair 195

Fig. 6. Specificity of the action of progesterone on myelination in lesioned peripheral nerves of wild type mice. The nerves were processed
for ultrastructural analysis 15 days after cryolesion (as described in Fig. 3 legend) after a series of four local applications of either the vehicle
(sesame oil), progesterone or progesterone blocker. (a) Male control nerve treated with the vehicle only. (b) Male nerve after trilostane treat-
ment (a progesterone synthesis inhibitor): the myelin sheaths are thinner than in controls. (c) Reversal of the effect of trilostane on myelin
sheaths by simultaneous administration of progesterone in male nerve, indicating that the decreased myelination is not due to the toxicity of
trilostane. (d) Nerve from a female ovariectomized mouse 10 days before cryolesion, treated with progesterone after the lesion. The myelin
sheaths are thicker than in male mice treated with progesterone. (e) Quantification of the effects of pregnenolone, progesterone and inhibitors
(mifepristone, RU486; trilostane; and trilostane plus progesterone) on thickness of myelin sheaths in male nerves, classified in increments of
five lamellae. The inset shows the average width of the myelin sheaths (that is, the number of lamellae). The data are obtained from the same
animals as used for data in Fig. 4, and are expressed as percentages of control (baseline). From 300 to 500 myelin sheaths were counted per
nerve. Progesterone-treated nerves were surrounded by thicker myelin sheaths than nerves from vehicle-treated mice. The action of pro-
gesterone on myelination was inhibited by drugs that inhibit the action of progesterone (modified from Koenig et al., 1995). During the
process of peripheral nerve regeneration, the thickness of the myelin sheaths is not related to the diameter of the axons. This absence of re-
lationship is maintained for life in Trembler hypomyelinated mice, both in regenerated and non-operated nerves.

pregnenolone to progesterone or mifepristone (RU486), a pro-

Effects of progesterone on myelination of injured axons in
the peripheral nervous system
Myelin originates from the Schwann cell surface membrane,
which forms a mesaxon that elongates and wraps spirally
around the axon. Further growth leads to a compact sheath
composed of concentrically arranged lamellae. The stimulatory
effect of progesterone on the remyelination (1) of regenerating
sciatic nerves in wild-type male mice (Fig. 6 a–c) and (2) of out-
grown neurites in DRG explants in culture (Fig. 2c,d,f) was first
demonstrated by Koenig et al. (1995). Progesterone increases the
rate of myelin synthesis and accelerates the initiation of myelin
formation in DRG neurone–Schwann cell cultures (Chan et al.,
1998). The application of progesterone to cryolesioned sciatic
nerves in mice stimulates thickening of the myelin sheaths
(Figs 6 and 7). Treatment with pregnenolone produces similar re-
sults (Koenig et al., 1995). Remyelination of cryolesioned sciatic
nerves is inhibited by trilostane, which blocks the conversion of
gestagen receptor blocker, by nearly 25 and 44%, respectively
(Figs 6 and 7). The inhibitory effect of trilostane is not due to
toxicity, since its effect on remyelination is reversed by the
simultaneous application of progesterone (Koenig et al., 1995
and Fig. 6c). At this stage of the remyelinating process, the
nerves recover their progesterone concentrations. In vitro,
RU486 does not influence myelin formation in DRG cultures,
but inhibits the stimulatory effect of the exogenous pro-
gesterone (Jung-Testas et al., 1999). Both exogenous pregnenol-
one and progesterone increase the neoformation of myelin
sheaths in lesioned peripheral nerves. A crucial question is:
which is the ‘active’ molecule? When the conversion of chol-
esterol to pregnenolone is blocked by aminoglutethimide, the
formation of myelin decreases (H. Koenig and B. Ferzaz, un-
published). Progesterone and its 5α-reduced metabolites, di-
hydroprogesterone and tetrahydroprogesterone, stimulate the
expression of genes encoding peripheral myelin proteins Po
196 H. L. Koenig et al.
(a) Mean number of myelin lamellae (b)
(as compared to the controls = 100)
Myelin sheaths thickness (%)
Treatment Control Progesterone
Male
Intact mice 12.9 ± 0.1 15.7 ± 0.1
Castrated mice 15.7 ± 0.5 17.4 ± 0.2
Female
Intact mice 15.8 ± 0.4 19.4 ± 1.0
Castrated mice 18.8 ± 0.1 21.4 ± 0.8
Fig. 7. Sexual dimorphism in the effects of progesterone on peripheral nerve remyelination. (a) Thickness of sciatic nerve myelin sheaths
(that is, number of lamellae) from intact and gonadectomized male and female mice, processed simultaneously for electron microscopy after
cryolesion and local treatment with the vehicle (control) or progesterone. (b) Thickness of sciatic nerve myelin sheaths from ovariectomized
females treated with progesterone, an inhibitor of progesterone synthesis (trilostane) or a progesterone receptor blocker (mifepristone, RU486).
Six to ten mice were analysed 15 days after they received four local applications of progesterone or progesterone blockers.
and PMP22; 3α,5α-tetrahydroprogesterone is more potent in
this respect than 5α-dihydroprogesterone and progesterone
(Melcangi et al., 1998). Therefore, the progesterone synthesized
in Schwann cells or the exogenous progesterone synthesized in
the sciatic nerve may serve as the parent molecule for several
potentially active steroids and steroid derivatives. Exogenous
progesterone also enhances the concentrations of Po and
PMP22 proteins in co-cultures of DRG neurones and Schwann
cells (Notterpeck et al., 1999) and stimulates the expression of
their gene promoters in cultured rat Schwann cells (Desarnaud
et al., 1998). These observations imply that progesterone binds
to progesterone receptors in the Schwann cell cytoplasm or nu-
cleus, although further investigation is required since pro-
gesterone receptor was not detected in mouse sciatic nerves
(I. Jung-Testas and H. Koenig, unpublished). Furthermore, in
progesterone receptor knock-out adult mice, the myelin sheaths
around sciatic nerve axons are the same as in wild-type mice
(Jung-Testas et al., 1999). Endogenous progesterone may acti-
vate the process of myelination through autocrine–paracrine
actions involving a limited number of progesterone receptors
(Koenig et al., 1995; Baulieu et al., 1996; Baulieu and
Schumacher, 1997). An alternative possibility is that the effect
of progesterone on myelination is mediated through GABAA
receptors (Melcangi et al., 1999). Finally, progesterone may
increase the transfer of axonally orthograde transported
phospholipids from axons to myelin (Droz et al., 1981; Toews
et al., 1988). Such a transfer would increase the rate of spiral
wrapping of the Schwann cell membrane, which constitutes the
myelin lamellae. This interpretation is sustained by two obser-
vations. First, remyelination begins 1 week or more after nerve
damage, when regenerating axons have already matured.
Second, in nerves treated for 7 days during the second week
after cryolesion, the thickness of myelin sheaths is similar to
500
RU486
400 TRIL
PROG
300
Castrated female
200
100
0
–100
–200
3–5 6–10 11–15 16–20 21–25 26–30 31–35 >35
Number of lamellae
that in nerves treated with progesterone for 15 days immedi-
ately after the lesion (P. Pelissier and H. Koenig, unpublished).
These observations indicate that progesterone is less, or not,
effective in stimulating myelination in axons while they are re-
generating. It remains to be determined whether progesterone
acts directly on Schwann cells or if its myelin-stimulating effect
is mediated through an axonal signal. An axonal signal is
known to be necessary for the initiation of the myelination
process (for review, see Salzer, 1995). The hypothetical mechan-
isms of the stimulatory effect of progesterone on remyelination
in lesioned nerves are shown (Fig. 5).
Progesterone in sexual dimorphism in peripheral
nerve myelination
Sexual dimorphism of reproductive steroid hormones is gen-
erally associated with sexual dimorphism in behaviour and
reproductive functions. Therefore, it is relevant to ask whether
the production of progesterone in peripheral nerves is also
sexually dimorphic. In rodents, the progesterone content of
the peripheral nerve is higher in females than that found in
males. In female mice, the concentrations of endogenous pro-
gesterone and pregnenolone differ among groups of nerves,
but are higher than in male sciatic nerves (M. Schumacher and
H. Koenig, unpublished). In female rats, the concentration of
progesterone in the sciatic nerves is higher in pro-oestrus (× 10)
than it is in oestrus (× 5), as compared with that in males
(Pelissier et al., 1996; M. Schumacher, unpublished). These obser-
vations prompt several questions. Does the higher progesterone
content in female nerves result in a faster rate of formation
of myelin in damaged nerve fibres in females than in males?
Does the administration of progesterone increase myelination
in lesioned nerves in males but not in females? As progesterone
 Role of progesterone in peripheral nerve repair 197

Progesterone

Neurotrophic THP
factors (NTF)
DHP
NTF-R
GABAA-R
Po, MBP, PR
PMP22 Po, PMP2...
Hyperpolarization
Myelin Gene activity Myelin Gene activity
wrapping wrapping

Spinal cord Axonal transport

motoneurone Axonal signal of phospholipids Axonal signal
or for myelination for myelination Target
dorsal root cell
ganglion
neurone Axonal signal Increased
transfer

Increa Increa
se in myelin wrapping se in myelin wrapping

Po, PMP22... Po, PMP22...

Schwann cell Schwann cell

Fig. 8. Possible mechanisms explaining the effects of progesterone on growth and maturation of the regenerating axons and consecutive
myelination in lesioned peripheral nerve. In vitro, progesterone may also directly stimulate neuronal soma, in addition to Schwann cells as it
does in in vivo experiments. PR: progesterone receptor; DHP: 5α-dihydroprogesterone; MBP: myelin basic protein; NTF-R: neurotrophic
factor receptor; PMP22: peripheral myelin protein 22; Po: protein Po; THP: 3α,5α-tetrahydroprogesterone; GABAA-R: GABAA receptor.

is mainly synthesized in the gonads, does castration influence
the myelination process of damaged peripheral nerve fibres in
a different way in males and in females?
In intact female mice, the mean number of myelin lamellae
formed around regenerating nerve fibres after cryolesion is
significantly higher (> 22%) than in regenerating nerves from
intact males (Fig. 7a). A similar difference is observed in ovari-
ectomized females (> 20%) as compared with castrated males
(Fig. 7a). In both genders, progesterone treatment enhanced
myelin width by approximately 22% and, in gonadectomized
animals, by > 11% in males and > 14% in females. These findings
indicate that the presence of gonadal hormones in the periph-
eral nerve and in the peripheral circulation stimulates myelin-
ation in damaged nerves. In intact and castrated progesterone-
treated animals of both sexes, the increase in mean thickness of
myelin is due to the increased number of thick myelin sheaths
made up of more than 20 lamellae. This increase of thick re-
generated myelin sheaths was identical in intact males and
females (> 125%), whereas it was different in gonadectomized
males (> 60%) and females (> 38%). However, in both intact
and ovariectomized females, the number of thick myelin sheaths
is higher than in males (intact and castrated). The faster rate of
remyelination in females is probably associated with the high
content of progesterone in female nerves (Pelissier et al., 1996)
and the higher concentrations of residual progesterone remain-
ing after cryolesion (M. Schumacher and H. Koenig, unpub-
lished). A similar difference is described in the rat hypoglossal
nerve (Yu, 1982) and in the male hamster facial nerve, in which
the axonal regeneration is 20% slower than in females (Kujawa
et al., 1991).
The question of the role of 5α-reduced metabolites of pro-
gesterone in remyelination of damaged peripheral nerves re-
mains to be analysed. In these nerves, activities of 5α-reductase
(an enzyme involved in the conversion of progesterone to 5α-
dihydroprogesterone) are high (Celotti et al., 1992). Furthermore,
small quantities of 3α,5α-tetrahydroprogesterone increase the
myelin protein Po expression in rat Schwann cell cultures
(Melcangi et al., 1998).
Initiation of myelination by progesterone in Trembler
mutant mice
In Trembler mice, the treatment of regenerating nerves with
progesterone results in the appearance of myelin lamellae at
198 H. L. Koenig et al.
day 7, after axonal maturation is nearly completed (Figs 3b and
4c). In Trembler cryolesioned untreated (control) nerves, myelin
is formed after day 10. Thus, it is likely that the accelerated for-
mation of the myelin sheaths in nerves of Trembler mice induced
by progesterone is a consequence of the faster maturation of the
lesioned axons. At later time intervals (after 12 and 20 days),
the increased number of myelinated fibres (Fig. 4c) and the in-
creased myelin width (Fig. 4d) in progesterone-treated nerves
may be the result of an increased rate of myelin synthesis or an
increase in the rate at which lamellae wrap around the axons.
Concluding remarks and future perspectives
The dual function of progesterone and its metabolites in the
regeneration of PNS nerves is well established. One function
of progestagens is to stimulate axonal growth, which entails,
sequentially, the maturation of the axons, their covering by
Schwann cells, the initiation of myelin formation from mes-
axons, and the elongation and spiral wrapping of Schwann cell
membranes inside the cytoplasm. A second function of pro-
gestagens is to accelerate the myelinating process directly.
Several intracellular pathways may be involved in the effects
of progesterone or its 5α-metabolites on axonal growth and
maturation and on the remyelination processes (Fig. 8).
In fact, nothing is known of the mechanisms involved in
progestagen-dependent PNS nerve regeneration. The mech-
anisms must involve progestagen-dependent functions in
both the damaged neurones and their associated Schwann cells,
possibly including changes in the gene transcription or protein
translation of proteins required for repair of the whole injured
system. Progesterone, or its derivatives, may act either directly
on the cells responsible for axonal regeneration or contribute to
the release of growth or myelin regulatory molecules at the site
of peripheral nerve injury. For both axonal growth and myelin-
ation, two questions arise: does progesterone act through its
receptor or through the GABAA receptors (Figs 5 and 8); and
what are the subsequent signalling pathways? In addition to
a study of myelin proteins and steroid enzymes stimulated
during nerve regeneration, research is needed on the effect
of progesterone on the enzymes involved in lipid synthesis in
both axon and Schwann cells. Axon membranes and myelin
contain high concentrations of lipids, the synthesis of which
may be stimulated by progesterone. Cholesterol transport
mechanisms are probably also important, since apolipoprotein
E is involved in axonal growth (Handelmann et al., 1992;
Nathan et al., 1994; Mahley et al., 1996) and apolipoprotein A–I
gene expression is increased in myelinating nerves (LeBlanc
et al., 1989). Other cells, such as macrophages and fibroblasts,
also secrete trophic substances, which might contribute to PNS
nerve regeneration, but to a lesser extent than steroids. If moto-
neurones and their axons are shown to be sensitive to pro-
gesterone or to its 5α-reduced metabolites, better understanding
of the molecular control of steroid modulation on axonal growth
and myelination is likely to lead to the development of thera-
pies for PNS diseases and spinal cord diseases.
This review is dedicated to Professor René Couteaux, for his 90th
birthday. The authors thank N. A Do Thi for providing new photo-
graphs of a common work (Koenig et al., 1995) and for expertise in
DRG cultures; B. Ferzaz for initiating the in vivo regeneration model
in our laboratory; M. Schumacher for contributing to the castration
of the mice; Prof. E. E. Baulieu for the first fellowship to W. H. Gong
(Société de Secours des Amis des Sciences) and J. Prémont for his
advice on the ‘GABA’ experiments. This work was supported by the
Université Bordeaux I and the Association Francaise contre les
Myopathies, who also provided grants to P. Pélissier and W. H. Gong.
At present, W. H. Gong is the recipient of a post-doctoral fellowship
from Institut pour la Recherche sur la Moelle Epinière. The authors
also thank J. Mallet for allowing H. Koenig and W. H. Gong to
pursue their research in his laboratory. H. Koenig is deeply grateful
to Prof. R. Couteaux for his permanent help and support. His enthusi-
asm and creativity in discussions were an exemple highly appreciated
and source of inspiration.
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