The present study provides evidence for the presence of hSOD1 aggregates with prion-like properties in the spinal ventral horn from a patient carrying the
hSOD1G127X mutation. The two different patient-derived aggregate seed preparations both transmitted a premature fatal motor neuron disease with hallmarks of human ALS: focal onset of progressive paralysis, loss of motor neurons and wasting. Concomitantly aggregation of hSOD1
G85R developed in the inoculated mice, which spread all along the neuraxis. The aggregates in end-stage
hSOD1G127X Tg mice had a strain A-like core structure and the induced aggregates displayed strain A patterns (Fig.
1c–e) [
7]. This suggests that the aggregation transmitted by the murine SOD1
G127X seed was templated. The human hSOD1
G127X seed also induced strain A hSOD1
G85R aggregation, but since the inducing aggregates could not be profiled, we cannot draw any firm conclusions regarding templating. Of note, induced aggregation in
hSOD1G85R Tg mice seems to replicate the structure of the inducing species: strain B aggregates from
hSOD1D90A Tg mice induce formation of strain B aggregates which do not arise spontaneously in
hSOD1G85R Tg mice (Fig.
1f) [
6,
7]. Further studies are needed to address the mechanisms by which the hSOD1 aggregation spreads in the CNS and causes the fatal neurotoxicity.
Aggregates of disease-relevant proteins have shown seeding effects in other models of neurodegenerative disease [
11,
12,
26,
28,
29]. The relevance of the findings for the diseases in humans has, however, been questioned [
13,
36]. In some cases, the induced pathology poorly mimics the modeled human disease, and the amounts of aggregates in the seeds and levels of expression of the disease-relevant proteins in the animals are considered unrealistically high. It is also suggested that the protein aggregation is related to specific vulnerabilities of subsets of neurons and might mainly occur cell autonomously in these.
In the current study, most of these concerns have been addressed. (1) As in humans, the fatal disease in
hSOD1G85R Tg mice has a middle-age onset, and demonstrates all the major hallmarks of human ALS. (2) The concentration of the hSOD1
G85R protein in the recipient mice is no higher than those of the endogenous murine SOD1 or hSOD1 in the human CNS [
8,
21]—although the proportion that is disordered is higher [
38,
39]. (3) The amounts of SOD1
G127X aggregates in the 1-µl inoculates were minute: 0.14–0.07 ng. This is around 500 times less than the amount of murine or human SOD1 in the same volume of spinal cord tissue from controls [
17,
22], and ~ 30 times less than the amount of aggregated hSOD1
G127X present in 1-µl (mg) ventral horn from patients carrying the
hSOD1G127X mutation [
19,
20]. Finally, the amounts of hSOD1
G85R aggregates in end-stage mice were > 30,000 times greater than the amounts of hSOD1
G127X aggregates that were inoculated. Thus, the seeding efficiency of the patient-derived hSOD1
G127X aggregates was extremely high, and they transmitted disease to the Tg mice at levels that were much lower than those found in motor areas of hSOD1
G127X patients. (4) The nature of the selective vulnerability of motor areas to SOD1-induced toxicity is still poorly understood. We have previously reported on two properties that would enhance the vulnerability to aggregation. Spinal cords from Tg model mice show a marked enrichment of disordered hSOD1 monomers [
38,
39], which are the necessary substrates for nucleation and growth of aggregate fibrils [
10,
16,
25]. Autophagy is found to be important for retarding hSOD1 aggregation in the transgenic models, and spinal motor areas in humans have apparently a low inherent such capacity [
35]. These two properties could conceivably enhance autonomous hSOD1 aggregation in cells compromised by other neurotoxic mechanisms, but would certainly also increase the risk of stochastic initiation and subsequent prion-like spread of hSOD1 aggregation preferentially in the motor areas. The patterns of spontaneous hSOD1 aggregation in the murine models support the latter possibility. Here, and previously, we have shown that strain A and B hSOD1 aggregate seeds initiate templated hSOD1 aggregations which spread from the site of inoculation, from which area also the initial motor symptoms derive [
7]. In
hSOD1G85R Tg mice that spontaneously develop disease or are inoculated with human or murine control seeds, the aggregation seems to initiate and spread from random positions along the spinal cord, with symptom onsets related to the segments with highest aggregate levels in the individual mice [
7].
We conclude that hSOD1 aggregate seeds prepared from spinal ventral horn from a patient carrying a hSOD1 mutation when inoculated in hSOD1-expressing mice initiated spreading hSOD1 aggregation concomitantly with fatal motor neuron disease. The potency of the seeds was extremely high, and disease was initiated in the Tg mice by levels of hSOD1G127X aggregates much lower than those found in the human motor system. Our results suggest that prion-like spread of hSOD1 aggregation could be the primary pathogenic mechanism, not only in hSOD1 Tg models, but also in hSOD1-induced ALS in humans.