Treatment with K6PC-5, a selective stimulator of SPHK1, ameliorates intestinal homeostasis in an animal model of Huntington's disease.

Emerging evidence indicates that Huntington's disease (HD) may be described as multi-organ pathology. In this context, we and others have contributed to demonstrate that the disease is characterized by an impairment of the homeostasis of gastro-intestinal (GI) tract. Sphingolipids represent a class of molecules involved in the regulation and maintenance of different tissues and organs including GI system. In this study, we investigated whether the alteration of Sphingosine-1-phosphate (S1P) metabolism, previously described in human HD brains and animal models, is also detectable peripherally in R6/2 HD mice. Our findings indicate, for the first time, that sphingolipid metabolism is perturbed early in the disease in the intestinal tract of HD mice and, its modulation by K6PC-5, a selective activator of S1P synthesis, preserved intestinal integrity and homeostasis. These results further support the evidence that modulation of sphingolipid pathways may represent a potential therapeutic option in HD and suggest that it has also the potential to counteract the peripheral disturbances which may usually complicate the management of the disease and affect patient's quality of life.

Curcumin dietary supplementation ameliorates disease phenotype in an animal model of Huntington's disease.

Huntington's disease (HD) has traditionally been described as a disorder purely of the brain; however, evidence indicates that peripheral abnormalities are also commonly seen. Among others, severe unintended body weight loss represents a prevalent and often debilitating feature of HD pathology, with no therapies available. It correlates with disease progression and significantly affects the quality of life of HD patients. Curcumin, a naturally occurring polyphenol with multiple therapeutic properties, has been validated to exert important beneficial effects under health conditions as well as in different pathological settings, including neurodegenerative and gastrointestinal (GI) disorders. Here, we investigated the potential therapeutic action that curcumin-supplemented diet may exert on central and peripheral dysfunctions in R6/2 mice, a well-characterized HD animal model which recapitulates some features of human pathology. Maintenance of normal motor function, protection from neuropathology and from GI dysfunction and preservation of GI emptying and conserved intestinal contractility, proved the beneficial role of life-long dietary curcumin in HD and corroborated the potential of the compound to be exploited to alleviate very debilitating symptoms associated with the disease.

Neuromedin U potentiates ADP- and epinephrine-induced human platelet activation.

Neuromedin U (NmU) is a pleiotropic hypothalamic neuropeptide involved in the gut-brain axis. It acts via both a Gαq/11-coupled receptor (NMUR1) and a Gαi-coupled receptor (NMUR2) in different cell types. Expression of both receptors was reported in platelets, but their significance for NmU signaling remains elusive. We studied the potential effects of NmU on human platelet activation. In platelet-rich plasma (PRP), NmU alone (up to 10µM) did not induce any measurable aggregation, but at nanomolar concentrations, it potentiated platelet aggregation by low (mean 0.47µM) ADP concentrations (from 25.9±3.6% to 74.8±2.7% maximal aggregation for ADP vs. ADP+NmU, 100nM, mean±SEM, n=13), accompanied by platelet P-selectin expression and intracellular calcium mobilization. Accordingly, platelet preincubation with NmU for 2min sensitized platelets for subsequent activation by ADP. When P2Y1 was inactivated by 50µM MRS2179, NmU comparably potentiated ADP-induced PRP aggregation, suggestive of cooperative activation with Gαi-coupled P2Y12. Likewise, NmU potentiated platelet aggregation by Gαi-operated epinephrine at subthreshold concentrations (99ng/ml, mean), but not that by Gαq-dependent serotonin (20µM). Platelet aggregation by NmU/epinephrine combination was fully inhibited by the Gαq inhibitor YM-254890 (1µM). qPCR detection and western blot analysis substantiated platelet expression of NMUR1 in different donors, a finding collectively complying with functionally relevant Gαq/11-mediated activation of platelet NMUR1 by NmU. Our findings advocate further studies on platelet sensitization by NmU, released during vascular activation and injury, to define its role as a modifier of platelet responsiveness to the physiological activation signals, operational in cardiovascular health and disease.

Motor phenotype is not associated with vascular dysfunction in symptomatic Huntington's disease transgenic R6/2 (160 CAG) mice.

Whereas Huntington's disease (HD) is unequivocally a neurological disorder, a critical mass of emerging studies highlights the occurrence of peripheral pathology like cardiovascular defects in both animal models and humans. The overt impairment in cardiac function is normally expected to be associated with peripheral vascular dysfunction, however whether this assumption is reasonable or not in HD is still unknown. In this study we functionally characterized the vascular system in R6/2 mouse model (line 160 CAG), which recapitulates several features of human pathology including cardiac disease. Vascular reactivity in different arterial districts was determined by wire myography in symptomatic R6/2 mice and age-matched wild type (WT) littermates. Disease stage was assessed by using well-validated behavioural tests like rotarod and horizontal ladder task. Surprisingly, no signs of vascular dysfunction were detectable in symptomatic mice and no link with motor phenotype was found.

MRI measures of corpus callosum iron and myelin in early Huntington's disease.

Increased iron in subcortical gray matter (GM) structures of patients with Huntington's disease (HD) has been suggested as a causal factor in neuronal degeneration. But how iron content is related to white matter (WM) changes in HD is still unknown. For example, it is not clear whether WM changes share the same physiopathology (i.e. iron accumulation) with GM or whether there is a different mechanism. The present study used MRI to examine iron content in premanifest gene carriers (PreHD, n = 25) and in early HD patients (n = 25) compared with healthy controls (n = 50). 3T MRI acquisitions included high resolution 3D T1, EPI sequences for diffusion tensor imaging (DTI) as an indirect measure of tissue integrity, and T2*-weighted gradient echo-planar imaging for MR-based relaxometry (R2*), which provides an indirect measure of ferritin/iron deposition in the brain. Myelin breakdown starts in the PreHD stage, but there is no difference in iron content values. Iron content reduction manifests later, in the early HD stage, in which we found a lower R2* parameter value in the isthmus. The WM iron reduction in HD is temporally well-defined (no iron differences in PreHD subjects and iron differences only in early HD patients). Iron level in callosal WM may be regarded as a marker of disease state, as iron does not differentiate PreHD subjects from controls but distinguishes between PreHD and HD.

Synergistic effects of BDNF and rehabilitative training on recovery after cervical spinal cord injury.

Promoting the rewiring of lesioned motor tracts following a spinal cord injury is a promising strategy to restore motor function. For instance, axonal collaterals may connect to spared, lesion-bridging neurons, thereby establishing a detour for descending signals and thus promoting functional recovery. In our rat model of cervical spinal cord injury, we attempted to promote targeted rewiring of the unilaterally injured corticospinal tract (CST) via the spared reticulospinal tract (RtST). To promote new connections between the two tracts in the brainstem, we administered viral vectors producing two neurotrophins. Brain-derived neurotrophic factor (BDNF), a known promotor of collateral growth, was expressed in the motor cortex, and neurotrophin 3 (NT-3), which has chemoattractive properties, was expressed in the reticular formation. Because rehabilitative training has proven to be beneficial in promoting functionally meaningful plasticity following injury, we added training in a skilled reaching task. Different neurotrophin or control treatments with or without training were evaluated. As hypothesized, improvements of motor performance with the injured forelimb following neurotrophin treatment alone were absent or modest compared to untreated controls. In contrast, we found a significant synergistic effect on performance when BDNF treatment was combined with training. The mechanism of this recovery remains unidentified, as histological analyses of CST and RtST collateral projections did not reveal differences among treatment groups. In conclusion, we demonstrate that following a cervical spinal lesion, rehabilitative training is necessary to translate effects of BDNF into functional recovery by mechanisms which are likely independent of collateral sprouting of the CST or RtST into the gray matter.

Mutant Huntingtin induces activation of the Bcl-2/adenovirus E1B 19-kDa interacting protein (BNip3).

Huntington's disease (HD) is a neurodegenerative disorder characterized by progressive neuronal death in the basal ganglia and cortex. Although increasing evidence supports a pivotal role of mitochondrial dysfunction in the death of patients' neurons, the molecular bases for mitochondrial impairment have not been elucidated. We provide the first evidence of an abnormal activation of the Bcl-2/adenovirus E1B 19-kDa interacting protein 3 (BNip3) in cells expressing mutant Huntingtin. In this study, we show an abnormal accumulation and dimerization of BNip3 in the mitochondria extracted from human HD muscle cells, HD model cell cultures and brain tissues from HD model mice. Importantly, we have shown that blocking BNip3 expression and dimerization restores normal mitochondrial potential in human HD muscle cells. Our data shed light on the molecular mechanisms underlying mitochondrial dysfunction in HD and point to BNip3 as a new potential target for neuroprotective therapy in HD.

The p85 regulatory subunit of PI3K mediates TSH-cAMP-PKA growth and survival signals.

Phosphatidylinositol 3-kinase (PI3K) is necessary for thyroid stimulating hormone (TSH)-induced cell cycle progression. To determine the molecular mechanism linking PI3K to TSH, we have identified a serine residue in p85alpha(PI3K) phosphorylated by protein kinase A (PKA) in vitro and in vivo. Expression of an alanine mutant (p85A) abolished cyclic AMP/TSH-induced cell cycle progression and was lethal in thyroid cells (FRTL-5). The aspartic version of the p85alpha(PI3K) (p85D) inhibited apoptosis following TSH withdrawal. The p85alpha(PI3K) wild type not the p85A bound PKA regulatory subunit RIIbeta in cells stimulated with cAMP or TSH. The binding of the aspartic version of p85alpha(PI3K) to RIIbeta was independent of cAMP or TSH stimulation. Similarly, binding of PI3K to p21Ras and activation of AKT, a downstream PI3K target, were severely impaired in cells expressing the p85A mutant. Finally, we found that the catalytic activity of PI3K was stimulated by TSH in cells expressing the wild-type p85alpha(PI3K) but not in cells expressing p85A. This latter mutant did not affect the epidermal growth factor-stimulated PI3K activity. We suggest that (1) TSH-cAMP-induced PKA phosphorylates p85alpha(PI3K) at serine 83, (2) phosphorylated p85alpha(PI3K) binds RIIbeta-PKA and targets PKAII to the membrane, and (3) PI3K activity and p21Ras binding to PI3K increase and activate PI3K downstream targets. This pathway is essential for the transmission of TSH-cAMP growth signals.