Gender effect on neurodegeneration and myelin markers in an animal model for multiple sclerosis.

BACKGROUND: Multiple sclerosis (MS) varies considerably in its incidence and progression in females and males. In spite of clinical evidence, relatively few studies have explored molecular mechanisms possibly involved in gender-related differences. The present study describes possible cellular- and molecular-involved markers which are differentially regulated in male and female rats and result in gender-dependent EAE evolution and progression. Attention was focused on markers of myelination (MBP and PDGFαR) and neuronal distress and/or damage (GABA synthesis enzymes, GAD65 and GAD67, NGF, BDNF and related receptors), in two CNS areas, i.e. spinal cord and cerebellum, which are respectively severely and mildly affected by inflammation and demyelination. Tissues were sampled during acute, relapse/remission and chronic phases and results were analysed by two-way ANOVA. RESULTS: 1. A strong gender-dependent difference in myelin (MBP) and myelin precursor (PDGFαR) marker mRNA expression levels is observed in control animals in the spinal cord, but not in the cerebellum. This is the only gender-dependent difference in the expression level of the indicated markers in healthy animals; 2. both PDGFαR and MBP mRNAs in the spinal cord and MBP in the cerebellum are down-regulated during EAE in gender-dependent manner; 3. in the cerebellum, the expression profile of neuron-associated markers (GAD65, GAD67) is characterized by a substantial down-regulation during the inflammatory phase of the disease, which does not differ between male and female rats (two-way ANOVA); 4. there is an up-regulation of NGF, trkA and p75 mRNA expression in the early phases of the disease (14 and 21 days post-immunization), which is not different between male and female. CONCLUSIONS: It is reported herein that the regulation of markers involved in demyelination and neuroprotection processes occurring during EAE, a well-established MS animal model, is gender- and time-dependent. These findings might contribute to gender- and phase disease-based therapy strategies.

Sex-dimorphic changes in neuroactive steroid levels after chronic experimental autoimmune encephalomyelitis.

Our previous observations have shown that neuroactive steroid levels in the brain are affected by acute experimental autoimmune encephalomyelitis (EAE) with sex and regional specificity (Giatti et al. 2010). To better understand the effect of EAE on neuroactive steroids, we have here assessed the levels of pregnenolone, progesterone and its derivatives (i.e. dihydroprogesterone, tetrahydroprogesterone and isopregnanolone), testosterone and its derivatives (dihydrotestosterone and 5alpha-androstane-3alpha, 17beta-diol) in different CNS regions of male and female rats affected by chronic EAE. Data obtained by liquid chromatography tandem mass spectrometry revealed that chronic EAE results in sex and regional specific alterations in the levels of neuroactive steroids in the brain, which are in many cases different to those produced by acute EAE. The specific changes in neuroactive steroid levels after chronic EAE may be of relevance to design new possible therapeutic strategies for the disease.

Acute experimental autoimmune encephalomyelitis induces sex dimorphic changes in neuroactive steroid levels.

Incidence, progression and severity of the multiple sclerosis, an inflammatory, demyelinating disease of the central nervous system (CNS) are affected in a sex-depending way. Physiological situations characterized by changes in sex steroid plasma levels, such as menstrual cycle, menopause or pregnancy, affect the disease course, suggesting that these molecules might exert a role in this disease. In order to understand better this possible relationship, we have here assessed the levels of neuroactive steroids present in different CNS regions of male and female rats affected by acute experimental autoimmune encephalomyelitis (EAE). In addition, we compared these levels with those present in plasma. Data obtained by liquid chromatography-tandem mass spectrometry indicate that the levels of neuroactive steroids show sex and regional differences in control and EAE nervous system and that a clear difference is also observed between CNS and plasma levels. In particular, among neuroactive steroids here considered, the levels of progesterone metabolites (i.e., dihydroprogesterone, tetrahydroprogesterone and isopregnanolone) and testosterone metabolites (i.e., dihydrotestosterone and 5alpha-androstane-3alpha17beta-diol), show sex dimorphic and region-specific changes in the CNS. Moreover, some changes observed in the CNS were not detected in plasma. These findings might represent an interesting background to design therapies and possibly sex-specific therapies for multiple sclerosis based on neuroactive steroids or synthetic ligands able to interact with classical and non-classical steroid receptors.

Gabapentin treatment improves motor coordination in a mice model of progressive ataxia.

No causal treatment of ataxias is available at the moment, and so symptomatic and disease-modifying therapies are regarded as a reliable possibility for this complex group of movement disorders. In order to explore possible pharmacological strategies aimed at interfering with ataxia development or progression, we used HCN1-/- mice. Mice carrying the deletion of the gene encoding for the voltage-dependent K-channel (HCN1-/-) have a normal basic motor function, but impaired learning of the motor skills that enable mice to balance on the rotating rod. In this study we showed that the motor coordination impairment observed in HCN1-/- mice is paralleled by a decline of GABA content in the cerebellum. Treatment with the GABA mimetic gabapentin at prenatal age prevents full development of the ataxia symptom. This result could have implications for possible therapeutic strategies aimed at more effective coupling with ataxia in several neurological diseases in which this symptom develops and is prominent over time. In view of its long-lasting effect, it could be also considered as a disease-modifying drug.

Hyperforin down-regulates effector function of activated T lymphocytes and shows efficacy against Th1-triggered CNS inflammatory-demyelinating disease.

Hyperforin (Hyp) is an active compound contained in the extract of Hypericum perforatum, well known for its antidepressant activity. However, Hyp has been found to possess several other biological properties, including inhibitory effects on tumor invasion, angiogenesis, and inflammation. In this paper, we show that treatment with Hyp inhibited IFN-gamma production, with down-regulation of T-box (T-bet; marker of Th1 gene expression) and up-regulation of GATA-3 (marker gene of Th2) on IL-2/PHA-activated T cells. In parallel, we showed a strong down-regulation of the chemokine receptor CXCR3 expression on activated T cells. The latter effect and the down-modulation of matrix metalloproteinase 9 expression may eventually lead to the inhibition of migratory capability and matrix traversal toward the chemoattractant CXCL10 by activated lymphocytes that we observed in vitro. The effect of Hyp was thus evaluated on an animal model of experimental allergic encephalomyelitis (EAE), a classic, Th1-mediated autoimmune disease of the CNS, and we observed that Hyp attenuates the severity of the disease symptoms significantly. Together, these properties qualify Hyp as a putative, therapeutic molecule for the treatment of autoimmune inflammatory disease sustained by Th1 cells, including EAE.

A molecular study of hippocampus in dogs with convulsion during canine distemper virus encephalitis.

In this study, we have investigated the expression of the nuclear transcription factor (c-Fos, NFkB), growth factors (nerve growth factor--NGF, brain-derived neurotrophic factor--BDNF), peptides (enkephalin, galanin) and glutamate transporter (AA 504-523 rat EAAC1) in 6 dogs sacrificed immediately after seizure attack during encephalomyelitis due to canine distemper virus (CDV) (as assessed by clinical examination, RT-PCR and viral RNA detection either in blood or brain tissue and CDV immunohistochemistry in brain slices). In all these CDV affected dogs, the observed neurological signs included untreatable seizures, leading to cluster seizure activity and status epilepticus. In the inter-ictal phase abnormal mentation, postural and gait deficits and sometimes involuntary movements such as myoclonus were recorded. The same investigation was carried out in 5 control dogs affected by different disorders, all characterized by the absence of seizures. Brains were dissected out immediately after euthanasia and fixed; sections collected from the dorsal hippocampus were processed for immunohistochemistry. By comparing hippocampus sections obtained from dog with and without seizure, the following regulations were observed. A strong up-regulation of glutamate transporter throughout the cell layers was found together with the onset of nuclear Fos and NFkB-IR in the pyramidal cell layer X. Among the investigated peptides, we observed a slight increase in enkephalinergic fibers and a strong up-regulation of mu-opioid receptors, whereas galanin-IR seemed to be weaker. Finally, both NGF and BDNF expression was strongly up-regulated. BDNF-IR was mainly localized in the apical dendrite in pyramidal neurons. To our knowledge, these data offer the first indication that molecular events described in experimental kindling also occur during spontaneous pathology in animal species sharing close similarities to human neuropathology.

Thyroid hormone and remyelination in adult central nervous system: a lesson from an inflammatory-demyelinating disease.

Re-myelination in the adult CNS has been demonstrated in different experimental models of demyelinating diseases. However, there is no clear evidence that re-myelination is effective in multiple sclerosis (MS), the most diffuse demyelinating disease. Moreover, chronic disabilities in MS are believed to be due to remyelination failure and consequent neuron damage and degeneration. Due to the presence of numerous oligodendrocyte precursors inside demyelination plaques, reasons for remyelination failure are unknown. In this paper, we reviewed data from embryonic development and in vitro studies supporting the primary role of thyroid hormone in oligodendrocyte maturation. We also reviewed personal data on the possibility of promoting myelination in chronic experimental allergic encephalomyelitis (EAE), a widely used experimental model of MS, by recruiting progenitors and channeling them into oligodendroglial lineage through the administration of thyroid hormone.

Cognitive deficit associated with cholinergic and nerve growth factor down-regulation in experimental allergic encephalomyelitis in rats.

Clinical symptoms in multiple sclerosis include cognitive dysfunction. Difficulties in learning and remembering new information represent the most common cognitive deficit and are associated with a general and progressive brain pathology. Possible pathogenetic mechanisms for neuronal damage such as neuroprotective strategies are under active investigation also in experimental allergic encephalomyelitis, the most widely used experimental model for multiple sclerosis. In this paper we demonstrate that a selective deficit in learning and memory performance, as investigated by the Morris water maze test, is a consistent feature in rat encephalomyelitis, which correlates with a decline in choline acetyltransferase activity and nerve growth factor mRNA level in cerebral cortex, hippocampus, and basal forebrain. Treatment aimed to restore acetylcholine content through chronic administration of selective acetylcholinesterase inhibitors (rivastigmine and donepezil) restores cognitive performance, choline acetyltransferase activity, and nerve growth factor mRNA expression.

Thyroid hormone administration enhances remyelination in chronic demyelinating inflammatory disease.

Chronic disabilities in multiple sclerosis are believed to be due to neuron damage and degeneration, which follow remyelination failure. Due to the presence of numerous oligodendrocyte precursors inside demyelination plaques, one reason for demyelination failure could be the inability of oligodendrocyte precursor cells to turn into myelinating oligodendrocytes. In this study, we show that thyroid hormone enhances and accelerates remyelination in an experimental model of chronic demyelination, i.e., experimental allergic encephalomyelitis in congenic female Dark Agouti rats immunized with complete guinea pig spinal cord. Thyroid hormone, when administered during the acute phase of the disease, increases expression of platelet-derived growth factor alpha receptor, restores normal levels of myelin basic protein mRNA and protein, and allows an early and morphologically competent reassembly of myelin sheaths. Moreover, thyroid hormone exerts a neuroprotective effect with respect to axonal pathology.

Stem cells and nervous tissue repair: from in vitro to in vivo.

Recent development in stem cell biology has indicated a new possible approach for the treatment of neurological diseases. However, in spite of tremendous hope generated, we are still on the way to understand if the use of stem cells to repair mature brain and spinal cord is a reliable possibility. In particular, we know very little on the in situ regulation of adult neural stem, and this also negatively impact on cell transplant possibilities. In this chapter we will discuss issues concerning the role and function of stem cells in neurological diseases, with regard to the impact of features of degenerating neurons and glial cells on in situ stem cells. Stem cell location and biology in the adult brain, brain host reaction to transplantation, neural stem cell reaction to experimental injuries and possibilities for exogenous regulation are the main topics discussed.

Neural stem cells and cholinergic neurons: regulation by immunolesion and treatment with mitogens, retinoic acid, and nerve growth factor.

Degenerative diseases represent a severe problem because of the very limited repair capability of the nervous system. To test the potential of using stem cells in the adult central nervous system as "brain-marrow" for repair purposes, several issues need to be clarified. We are exploring the possibility of influencing, in vivo, proliferation, migration, and phenotype lineage of stem cells in the brain of adult animals with selective neural lesions by exogenous administration (alone or in combination) of hormones, cytokines, and neurotrophins. Lesion of the cholinergic system in the basal forebrain was induced in rats by the immunotoxin 192IgG-saporin. Alzet osmotic minipumps for chronic release (over a period of 14 days) of mitogens [epidermal growth factor (EGF) or basic fibroblast growth factor (bFGF)] were implanted in animals with behavioral and biochemical cholinergic defect and connected to an intracerebroventricular catheter. After 14 days of delivery, these pumps were replaced by others delivering nerve growth factor (NGF) for an additional 14 days. At the same time, retinoic acid was added to the rats' food pellets for one month. Whereas the lesion decreased proliferative activity, EGF and bFGF both increased the number of proliferating cells in the subventricular zone in lesioned and nonlesioned animals. These results are indicated by the widespread distribution of BrdUrd-positive nuclei in the forebrain, including in the cholinergic area. Performance in the water maze test was improved in these animals and choline acetyltransferase activity in the hippocampus was increased. These results suggest that pharmacological control of endogenous neural stem cells can provide an additional opportunity for brain repair. These studies also offer useful information for improving integration of transplanted cells into the mature brain.