Institutionen för neurovetenskap / Department of Neuroscience
Abstract
The capacity of motoneurons to survive and regenerate new axons after
injuries such as axotomy depends on a great variety of parameters. After
sciatic nerve transection (SNT) in adult rats and to some extent after a
ventral funiculus lesion (VFL), many cells survive and regeneration is
highly prioritated. After SNT in neonatal rats or ventral root avulsion
(VRA) in adult rats, on the other hand, a large proportion of the
lesioned cells die and overall chances of regeneration are small.
Differences in protein expression patterns of trophic factors, their
receptors and receptors to extracellular matrix molecules, integrins,
were studied in motoneurons and sear tissue after the mentioned types of
injury. The effect of infiltrating inflammatory cells and substances
produced by such cells was studied on rat motoneurons in vivo and in cell
cultures, with the aim to determine some of the factors that have an
influence on the outcome of the injured motoneuron.
In all lesion models the expression of mRNA encoding growth-associated
protein (GAP-43) was highly upregulated. Alpha-CGRP was downregulated in
response to a SNT in newborn rats, but was upregulated in two week old
and in adult rats. GAP-43 and CGRP mRNA expressions in spinal motoneurons
are regulated, at least in part, by target-derived factors like CNTF and
bFGF and may be nerve cell-derived substances that are involved in
sprouting and regulation of neuromuscular junctions during development
and regeneration.
The trophic factors glial cell-line derived neurotrophic factor (GDNF),
brain-derived neurotrophic factor (BDNF), neurotrophin-3 (NT-3), ciliary
neurotrophic factor (CNTF), leukemia inhibitory factor (LIF) and
insulin-like growth factors (IGF-1 and -2) have been shown to be crucial
for survival and regeneration of spinal motoneurons after various kinds
of axotomy and in vitro. The mRNA expression in transected sciatic nerve
was correlated to the regulation of their receptor subunits in spinal
motoneurons and compared in the different lesion models. GDNF receptor
alpha (GFR[alpha]1), c-RET and LIF receptor (LIFR) mRNAs were extensively
upregulated after axotomy, with an earlier response after avulsion and
highly prioritated expression of GFR[alpha]1 in surviving motoneurons.
Levels of CNTF receptor alpha (CNTFR[alpha]) and the BDNF high affinity
receptor trkB were rather unaffected. The largest difference in response
between SNT and VRA in the adult rat was found for the NT-3 receptor trkC
and membrane glycoprotein 130 (gpl30) mRNAs. Thus, after SNT there were
only minor changes, while VRA induced a profound down-regulation.
IGF-1 is available for lesioned motoneurons both after peripheral and
central axonal lesions, while significant IGF-2 expression was restricted
to denervated nerve. Also the regulation of IGF binding proteins (IGFBPs)
are important. Thus, IGFBP-4 and IGFBP-5 were dominant in peripheral
nerve, while IGFBP-2 and IGFBP-5 were preferentially found in the CNS.
IGFBP-6 mRNA was strongly upregulated in spinal motoneurons and may be of
special relevance for the severed cells, probably in relation to the
IGF-2 expression in denervated nerve.
Both integrin subunits [alpha]7 and ß1 were highly upregulated in spinal
motoneurons after SNT, VRA and VFL and, in addition, integrin [alpha]6,
which is not normally detectable. The levels of integrin 0 remained
elevated longer after SNT than after VRA and VFL whereas integrin
[alpha]6 displayed the opposite pattern. Interestingly, at 21 and 42 days
after VFL, some motoneurons once again upregulated [alpha]7, possibly
indicating a regeneration through the scar and contact with ventral
roots.
Also taken in consideration was the lesion-accompanied local inflammatory
response. The traditional view has been that the relative immune
privilege of the CNS serves to protect delicate neuronal networks from
damage by immune reactions, but herein is demonstrated that axotomy
induces an upregulation of MHC I and ß2-microglobulin mRNA in
motoneurons, indicating an active participation in the immune response
after injury.
Experimental autoimmune encephalomyelitis (EAE) induced by active
immunization with an encephalitogenic MBP peptide led to a robust
survival promoting effect on avulsed motoneurons in spite of a very
intense inflammatory reaction with high levels of pro-inflammatory
cytokines in the lesioned segments. Furthermore, the expression of the
neurotrophic factors BDNF, NT-3 and GDNF by encephalitogenic T and NK
cells, may constitute an important mechanism for neuronal protection in
CNS inflammation and is supported by the fact that interferon-[gamma]
(IFN-[gamma]) or tumor necrosis factor-[alpha] (TNF-[alpha]) treatment
increased death of motoneurons only in cultures deprived of neurotrophic
support.
In conclusion, this thesis demonstrates a complex regulation of a network
of substances that are decisive for the regenerative potential of
motoneurons after mechanical nerve trauma