**8. ALS and riluzole**

gamma [5]. Treatment with PPAR gamma agonists decreases mRNA and protein levels of betacatenin in 3T3L1 adipocytes [1]. TZDs induce a reduction in the levels of cytoplasmic betacatenin in hepatocytes [3]. PPAR gamma suppresses Wnt/beta-catenin pathway during

**6. Deactivation of the Wnt/beta-catenin pathway induces activation of**

Inhibition of Wnt/beta-catenin pathway leads to an increase in transcription of PPAR gamma. Activation of the Wnt/beta-catenin signaling leads to osteogenesis, but not to adipogenesis. The canonical Wnt/beta-catenin-PPAR gamma system regulates the molecular switching of osteablastogenesis versus adipogenesis [6]. Wnt signaling maintains preadipocytes in an undifferentiated state through inhibition of both adipogenic transcription factors C/EBP alpha and PPAR gamma. Deactivation of Wnt/beta catenin pathway and activation of PPAR gamma are observed in ARVD [4, 31]. Taken together, these studies suggest that the canonical Wnt/ beta-catenin signaling downregulates PPAR gamma expression, inhibition of Wnt/betacatenin signaling upregulates PPAR gamma expression and PPAR gamma agonists inhibit the

The canonical Wnt/beta-catenin signaling is involved in numerous NDs, particularly in ALS. Several studies have shown that this pathway is upregulated in motor neurons of ASL model mice [32–35]. In the spinal cord of SOD1(G93A) ALS transgenic mice, expression of Wnt2, Wnt7a and GSK-3beta has been determined [32]. Both Wnt2, Wnt7a mRNA and protein in the spinal cord of ALS mice have been found to be upregulated when compared with wild type. The immune-reactivity of Wnt2 and Wnt7a is strong in ALS adult transgenic mice, whereas it is weak in wild-type mice. Neurodegeneration upregulates the expression of Wnt2 and Wnt7a in the spinal cord of ALS mice, which in turn activates Wnt signaling and inhibits GSK-3beta activity in ALS adult transgenic mice. Expression of Wnt3a, beta-catenin and Cyclin D1, three key molecules of the Wnt/beta-catenin signaling, have been determined in the adult spinal cord of SOD1(G93A) ALS transgenic mice at different stages [33]. It has been found that mRNA and protein of Wnt3a and Cyclin D1 in the spinal cord of the ALS mice are upregulated compared with wild-type mice. Moreover, beta-catenin translocates from the cell membrane to the nucleus and subsequently activated transcription of the target gene Cyclin D1. Wnt3a, beta-catenin and Cyclin D1 are also expressed in both neurons and astrocytes. For the authors, these findings suggest that neurodegeneration activates the Wnt/beta-catenin pathway, in the spinal cord of adult ALS transgenic mice. Changes in Wnt5a and Fzd2 expression in the spinal cord of SOD1(G93A) transgenic mice (ALS), SOD1(G93A) transfected NSC-34 cells and primary cultures of astrocytes from SOD1(G93A) transgenic mice have been observed [35]. Expression of Wnt1 and Fzd1 has been found to be increased in the spinal cords of SOD1G93A

adipogenesis [2].

260 Update on Amyotrophic Lateral Sclerosis

**PPAR gamma**

canonical Wnt/beta-catenin pathway.

**7. ALS and Wnt/beta-catenin pathway**

Today, no really efficient treatment exists for ALS [38, 39]. However, riluzole has been approved for the treatment of ALS in most countries and is tested in people based on results supporting a role of glutamate toxicity in ALS. Riluzole has numerous pharmacodynamics properties, i.e., presynaptic inhibition of the glutamate release, inhibition of G-proteindependent processes, modulation of N-methyl-D-aspartate ionotropic receptor and blockade of the voltage-gated sodium channel, etc. [39]. Two trials [12, 13] have demonstrated the weak efficacy of riluzole in ALS with prolongation of median survival by 2 to 3 months and safety of riluzole. Thus, riluzole appears to slow the progression of ALS, and may improve survival in patients with disease of bulbar onset [12]. Riluzole is well tolerated and lengthens survival of patients with ALS [13]. Two other studies have led to almost the same conclusions [14, 15]. The FDA-approved drug, riluzole, 100 mg daily is reasonably safe and probably prolongs median survival by about 2 to 3 months in patients with ALS.

Importantly, riluzole has been found to be an enhancer of the Wnt/beta-catenin signaling in melanoma [40]. For the authors, treating melanoma cells with riluzole in vitro enhances the ability of WNT3A to regulate gene expression, promote pigmentation and decrease cell proliferation. Like WNT3A, riluzole decreases metastases in a mouse melanoma model. Moreover, riluzole enhances Wnt/beta-catenin signaling in the primary screen both in HT22 neuronal cells and in adult hippocampal progenitor cells [40]. As the Wnt/beta-catenin pathway is upregulated, at least in genetic ALS mice [32–35], this can partly explain poor results in trials testing riluzole in ALS as shown previously [12–15]. Lithium, an activator of the Wnt/beta catenin signaling, has also been evaluated as a treatment for ALS [41]. Surpris‐ ingly, in ALS patients treated with lithium, the disease progression has been shown to be markedly attenuated. In the genetic ALS G93A mouse model, there is a marked neuroprotec‐ tion induced by lithium, which delayed disease onset and duration and augmented the life span. The use of the enhancer Wnt/beta-catenin lithium can be discussed in ALS in which the Wnt/beta-catenin pathway has been shown to be upregulated in several animal studies [32– 35]. GSK-3beta-inhibitor lithium chloride enhances activation of the canonical Wnt signaling [42–44]. Lithium activates downstream components of the Wnt signaling pathway in vivo, leading to an increase of the beta-catenin protein. This pathway is implicated in the patho‐ physiology and treatment of bipolar disorder [45, 46]. Riluzole reduces symptoms of obsessivecompulsive disorder, unipolar and bipolar depression and generalized anxiety disorder [47]. This is not surprising due to the fact that the Wnt/beta-catenin pathway is downregulated in bipolar syndrome [8] and that like lithium, riluzole is an enhancer of Wnt/beta-catenin signaling.

#### **9. ALS and PPAR gamma**

In ALS, expression of PPAR gamma (mARN and protein) has not been precisely investigated in neurons. However, the upregulation of Wnt/beta-catenin signaling observed in ALS suggests that PPAR gamma might be downregulated due to the fact that these two systems generally operate in the opposite way [1–3, 5]. Neuroinflammation is a common pathological feature in NDs, particularly in ALS. PPAR gamma may be a key regulator of neuroinflamma‐ tion. PPAR gamma inhibits NF-kappaB-mediated inflammatory signaling at multiple sites [48]. PPAR gamma might be a relevant regulator of neuroinflammation and possibly a new target for the development of therapeutic strategies for ALS. A potentially therapeutic pathway in ALS may be the activation by PPAR gamma agonists due to their ability to block neuropa‐ thological damages caused by inflammation [49]. The neuroprotective effect of pioglitazone has been demonstrated in G93A SOD1 transgenic mouse model of ALS and shows a significant increase in their survival. Pioglitazone protects motor neurons against p38-mediated neuronal death and NF-kappaB-mediated glial inflammation via a PPAR gamma-independent mecha‐ nism [50]. In ALS, PPAR gamma controls natural protective mechanisms against lipid peroxidation [51]. PPAR gamma-driven transcription selectively increases in the spinal cord of hSOD1G93A mice. This is correlated with the upregulation of lipid detoxification enzymes such as the lipoprotein lipase and glutathione S-transferase alpha-2, implied in scavenging lipid peroxidation by-products. Anticipation of protective reactions by pharmacological PPAR gamma modulation of the transcriptional activity attenuates neurodegeneration induced by lipid peroxidation. PPAR gamma activation is neuroprotective in a Drosophila model of ALS [52]. This Drosophila model of ALS based on TDP-43 recapitulates several aspects of ALS pathophysiology. Pioglitazone rescues TDP-43-dependent locomotor dysfunction in motor neurons and glia. PPAR gamma activation in neurons and glia is partially neuroprotective and restores metabolic alterations in ALS. Superoxide dismutase (SOD1)-G93A transgenic mice benefit from oral treatment with the PPAR gamma agonist pioglitazone [53]. Pioglitazonetreated transgenic mice reveal improved muscle strength and body weight, exhibit a delayed disease onset and survive significantly longer than non-treated SOD1-G93A mice. Pioglita‐ zone-induced neuroprotection of motor neurons of the spinal cord is complete at day 90. There is also preservation of the median fiber diameter of the quadriceps muscle, indicating a morphological and functional protection of motor neurons induced by pioglitazone. However, in a phase II double-blind controlled clinical trial, the PPAR gamma agonist pioglitazone in combination with riluzole does not increase survival in ALS patients [54].

PPAR gamma coactivator-1alpha (PGC-1alpha) is a transcriptional coactivator that works together with the transcription factor PPAR gamma in the regulation of mitochondrial biogenesis. PGC-1alpha plays a role in several neurodegenerative pathologies [26]. PGC-1al‐ pha protects neurons and alters disease progression in a PGC-1alpha transgenic mice crossed with SOD1 mutant G93A DL mice [55]. In these mice, the progression of the disease has been shown to be significantly slower. There is also a markedly improved performance on the rotarod test associated with an improved motor activity with a decreased loss of motor neurons and less degeneration of neuromuscular junctions. By using a double transgenic mouse model where PGC-1alpha is over-expressed in a SOD1 transgenic mouse (TgSOD1-G93A/ PGC-1alpha), it has been found that motor function and survival are improved [56]. This is accompanied by a reduction of motor neuron loss, a restoration of mitochondrial electron transport chain activities and an inhibition of stress signaling in the spinal cord. Thus, in the double-transgenic mice, there are improved motor performance, slowed ALS progression, decreased weight loss, and reduced motor neuronal death. Survival and disease improvement are greater in higher-expressing PGC-1alpha mice. Therefore, PPAR gamma is a possible target for ALS as it functions as a transcription factor that interacts with PGC-1alpha. Elevated PGC-1alpha activity sustains mitochondrial biogenesis and muscle function without extend‐ ing survival in a mouse model of inherited ALS [57]. Increasing PGC-1alpha activity in muscles represents an attractive therapy for maintaining muscle function during the progression of ALS.
