Wnt and lithium: a common destiny in the therapy of nervous system pathologies?

Wnt signaling is required for neurogenesis, the fate of neural progenitors, the formation of neuronal circuits during development, neuron positioning and polarization, axon and dendrite development and finally for synaptogenesis. This signaling pathway is also implicated in the generation and differentiation of glial cells. In this review, we describe the mechanisms of action of Wnt signaling pathways and their implication in the development and correct functioning of the nervous system. We also illustrate how a dysregulated Wnt pathway could lead to psychiatric, neurodegenerative and demyelinating pathologies. Lithium, used for the treatment of bipolar disease, inhibits GSK3ß, a central enzyme of the Wnt/ß-catenin pathway. Thus, lithium could, to some extent, mimic Wnt pathway. We highlight the possible dialogue between lithium therapy and modulation of Wnt pathway in the treatment of the diseases of the nervous system.

Bacterial lipopolysaccharide induces type 2 deiodinase in cultured rat astrocytes.

In the brain, 3,5,3'-triiodothyronine, which binds to the thyroid hormone receptor with high affinity, is locally generated from thyroxine by type 2 iodothyronine deiodinase (D2) expressed mainly in astrocytes and tanycytes. We have investigated the effects of bacterial lipopolysaccharide (LPS) on D2 in cultured rat astrocytes. LPS induced D2 activity with a lag-time of 4-8 h and a maximum at 24 h. LPS also promoted D2 mRNA accumulation. Glucocorticoids enhanced both the basal and LPS-stimulated D2 activity and mRNA accumulation. These glucocorticoid effects were blocked by the glucocorticoid receptor antagonist RU486. Our results obtained with different specific signaling pathway inhibitors indicated that D2 induction by LPS required ERK and p38-MAPK signaling pathways. NF-κB inhibitor sulfasalazine blocked the effects of LPS on both D2 activity and mRNA accumulation. Hence, D2 induction by LPS appeared to implicate NF-κB pathway in astrocytes. NF-κB responsiveness of the rat dio2 gene was studied in astrocytes with dio2 5'-flanking region promoter assays. The long form of the dio2 promoter was transactivated by NF-κB. CCAAT/enhancer-binding protein ß, which is upregulated by LPS in astrocytes, increased the transcriptional activity of the dio2 promoter in its long or truncated forms containing CCAATs. Our observations, which demonstrate D2 induction by LPS in astrocytes and specify some characteristics of D2 induction mechanism, support the possible implication of brain D2 in adaptative responses to an infectious stress.

Implication of type 3 deiodinase induction in zebrafish fin regeneration.

Thyroid hormones are critical determinants of cellular differentiation. We used the zebrafish model to evaluate the involvement of thyroid hormones in regeneration processes after caudal fin amputation. We examined early events following fin amputation, i.e., blastema formation and nerve repair by growth cone formation. Here, we show that the abolition of thyroid gland activity by methimazole treatment had no effect on blastema formation, but slowed growth cone formation of the lateral line. Conversely, the addition of exogenous thyroid hormones enhanced growth cone formation without affecting blastema formation. However, amputation triggered a strong induction in the blastema of type 3 deiodinase mRNA and enzymatic activity, which degrades thyroid hormone (TH). We therefore blocked deiodinase activity with iopanoic acid (IOP) and saw a reduction in blastema formation, suggesting that local degradation of TH is permissive for cell proliferation in the blastema. The effect of IOP on the blastema required endogenous or exogenous TH. Our findings support a model in which local degradation of TH by type 3 deiodinase is permissive for epimorphic regeneration.

Role of H2O2 in RET/PTC1 chromosomal rearrangement produced by ionizing radiation in human thyroid cells.

During childhood, the thyroid gland is one of the most sensitive organs to the carcinogenetic effects of ionizing radiation that may lead to papillary thyroid carcinoma (PTC) associated with RET/PTC oncogene rearrangement. Exposure to ionizing radiation induces a transient "oxidative burst" through radiolysis of water, which can cause DNA damage and mediates part of the radiation effects. H(2)O(2) is a potent DNA-damaging agent that induces DNA double-strand breaks, and consequently, chromosomal aberrations. Irradiation by 5 Gy X-ray increased extracellular H(2)O(2). Therefore, we investigated the implication of H(2)O(2) in the generation of RET/PTC1 rearrangement after X-ray exposure. We developed a highly specific and sensitive nested reverse transcription-PCR method. By using the human thyroid cell line HTori-3, previously found to produce RET/PTC1 after gamma-irradiation, we showed that H(2)O(2), generated during a 5 Gy X-ray irradiation, causes DNA double-strand breaks and contributes to RET/PTC1 formation. Pretreatment of cells with catalase, a scavenger of H(2)O(2), significantly decreased RET/PTC1 rearrangement formation. Finally, RET/PTC chromosomal rearrangement was detected in HTori-3.1 cells after exposure of cells to H(2)O(2) (25 micromol/L), at a dose that did not affect the cell viability. This study shows for the first time that H(2)O(2) is able to cause RET/PTC1 rearrangement in thyroid cells and consequently highlights that oxidative stress could be responsible for the occurrence of RET/PTC1 rearrangement found in thyroid lesions even in the absence of radiation exposure.

Oxidative stress regulates type 3 deiodinase and type 2 deiodinase in cultured rat astrocytes.

Type 2 deiodinase (D2) and type 3 deiodinase (D3) locally achieve the determination of the concentration of T3, which binds to the thyroid hormone receptor with high affinity. D2 converts T4 into T3, and D3 degrades T4 and T3. Neurons take up T3 released by astrocytes, the main cerebral site for the D2 expression. Because oxidative stress is believed to be involved in several neurological disorders, we explored the effects of oxidative stress on D3 and D2 in primary culture of rat astrocytes. H2O2 (250 microm) increased D3 activity with maximal effects around 8 h. Stimulation of D3 activity by H2O2 was synergistic with T4, phorbol ester, and also cAMP. H2O2 (250 microm) did not affect basal D2 activity but inhibited the stimulation of D2 activity by cAMP and factors implicating cAMP-independent pathways in astrocytes, TSH, and phorbol ester. N-Acetyl cysteine and selenium repletion, which respectively increase intracellular glutathione and glutathione peroxidase, inhibited D2 and D3 regulation by H2O2, whereas L-buthionine sulfoximine, which decreases intracellular glutathione, mimicked H2O2 effects. Oxidative stress up-regulated D3 and inhibited cAMP-stimulated D2 by transcriptional mechanisms. A decrease in cAMP by oxidative stress could contribute to the inhibition of cAMP-stimulated D2. Using specific inhibitors of signaling pathways, we show that the ERK pathway was required in D2 and D3 regulation by oxidative stress and that the p38 MAPK pathway was implicated in H2O2-induced D3. We suggest that the expected decrease in T3 might modulate the cellular injury of oxidative stress in some pathological brain conditions.

Hypoxia stabilizes type 2 deiodinase activity in rat astrocytes.

T(4) activation into T(3) is catalyzed by type 2 deiodinase (D2) in the brain. The rapid induction of D2 in astrocytes by transient brain ischemia has prompted us to explore the effects of hypoxia on D2 in cultures of astrocytes. Hypoxia (2.5% O(2)) of cultured astrocytes increased D2 activity, alone or in association with agents stimulating the cAMP pathway. Hypoxia had no effect on D2 mRNA accumulation. Cycloheximide did not block the effect of hypoxia on D2 activity and D2 half-life was enhanced under hypoxia demonstrating a posttranslational action of hypoxia. Furthermore, the D2 activity increase by hypoxia was not additive with the increase promoted by the proteasome inhibitor carbobenzoxy-L-leucyl-L-leucyl-L-leucinal (MG132). This strongly suggests that hypoxia leads to stabilization of D2 by slowing its degradation by the proteasome pathway. Hypoxia, in contrast to MG132, did not block the T(4)-induced D2 inactivation. A contribution of prolyl hydroxylase to the hypoxia effects on D2 was also suggested on the basis of increased D2 activity after addition of different prolyl hydroxylase inhibitors (cobalt chloride, desferrioxamine, dimethyloxalylglycine, dimethylsuccinate). Specific inhibitors of ERK, p38 MAPK, or phosphatidylinositol 3-kinase pathways were without any effect on hypoxia-increased D2 activity, eliminating their role in the effects of hypoxia. Interestingly, diphenyleneiodonium, an inhibitor of nicotinamide adenine dinucleotide phosphate oxidase inhibited the hypoxia-increased D2 indicating a role for some reactive oxygen species in the mechanism of D2 increase. Further studies are required to clarify the precise molecular mechanisms involved in the D2 stabilization by hypoxia.

Cloning and characterization of a novel isoform of iodotyrosine dehalogenase 1 (DEHAL1) DEHAL1C from human thyroid: comparisons with DEHAL1 and DEHAL1B.

The human iodotyrosine dehalogenase 1 (DEHAL1) gene is composed of six exons. Two isoforms (DEHAL1 and DEHAL1B) have been published in GenBank, both of which have a nitroreductase domain and arise from differential splicing in exon 5. We recently showed that the DEHAL1 isoform is a transmembrane protein that efficiently catalyzes the NADPH-dependent deiodination of mono (L-MIT) and diiodotyrosine (L-DIT) in human embryonic kidney-293 (HEK293) cells. In the present study, we establish the existence of a new transcript, DEHAL1C, in the human thyroid with a terminal exon that lacks in the DEHAL1 transcript. This exon is the complete exon 5, which is spliced in the DEHAL1B mRNA variant. These two variants encode proteins with differing C-terminal domains. Using quantitative reverse transcription polymerase chain reaction, we found that the expression of the mRNA of DEHAL1C and DEHAL1B was lower than that of DEHAL1 mRNA in the thyroid. We also observed that human DEHAL1B and DEHAL1C proteins are rapidly degraded in stably transfected HEK293 cells, unlike the DEHAL1 protein, and that exposure to the proteasome inhibitor MG132 resulted in accumulation of these proteins that was markedly time- and concentration-dependent. These findings show that the cytoplasmic tail could play a role in the stability of the protein.

Thyroid hormone deiodinases in the central and peripheral nervous system.

Thyroid hormones play a critical role in development and functioning of the nervous system. Deiodinases (type 2 [D2] and type 3 [D3]) contribute to the control of thyroid hormone action in the nervous system by regulating the local concentrations of triiodothyronine (T(3)), the main active thyroid hormone. Most brain T(3) is indeed locally formed by deiodination of thyroxine (T(4)). This reaction is catalyzed by D2 expressed in astrocytes throughout the brain and in tanycytes in the mediobasal hypothalamus. D3, which inactivates both T(4) and T(3), is mainly expressed in neurons also throughout the brain, with high expression in hippocampus and pyriform cortex. The regulation of deiodinases by many factors in addition to the thyroid hormones indicate that their role is not limited to mitigate the fluctuations in plasma T(4) and T(3). In contrast to the brain, deiodinases are not expressed in the adult peripheral nerve. Nerve lesions induce D2 in peripheral nerve sheaths and D3 in the endoneurial compartment containing Schwann cells. On the basis of available data summarized in this review, D2 and D3 clearly contribute to determine T(3) concentrations depending on the area of the nervous system, the state of development, and the pathophysiologic conditions.

Induction of type 2 iodothyronine deiodinase in astrocytes after transient focal cerebral ischemia in the rat.

This study investigated the expression of deiodinases of thyroid hormones in the rat brain after transient occlusion of the middle cerebral artery. The activity of type 2 deiodinase (D2), which catalyzes the deiodination of thyroxine into the more active thyroid hormone 3,5,3'-triiodothyronine, was strongly increased by cerebral ischemia at 6 and 24 hours in the striatum and at 24 hours in the cerebral cortex. The activity of type 3 deiodinase, which catalyzes the inactivation of thyroid hormones, was not affected by ischemia. In situ hybridization showed, as soon as 6 hours, an upregulation of the expression of D2 mRNA in the ipsilateral striatum, which disappeared at 24 hours. In the ipsilateral cortex, the induction of D2 mRNA started at 6 hours, was increased at 24 hours and finally declined at 72 hours. These results were confirmed by reverse transcription-PCR for D2 mRNA in the striatum and cerebral cortex. The upregulation of D2 mRNA after ischemia was mainly localized in astrocytic cell bodies. These results show that D2 is rapidly induced in astrocytes after ischemic stroke. Future work will include the exploration of the role of the upregulation of this enzyme, responsible for local 3,5,3'-triiodothyronine production as a neuroprotective mechanism in the brain.

MAP kinase activation by fluoxetine and its relation to gene expression in cultured rat astrocytes.

Chronic treatments with antidepressants active on major depressive disorders influence pathways involved in cell survival and plasticity. As astrocytes seem to play a key role in the protection of brain cells, we investigated in these cells the rapid effects of the antidepressant fluoxetine (Prozac) on signaling cascades and gene induction, which probably play a role in neuroprotection. We show here that fluoxetine alone activates the extracellular signal-regulated-protein kinase (Erk) and p38 mitogen-associated protein (MAP) kinase cascades. RT-PCR revealed that genes, modulated in brain by long-term fluoxetine treatment, are rapidly induced by fluoxetine in cultured astrocytes: brain-derived nerve factor (BDNF) and its receptors, glial-derived nerve factor (GDNF) and deiodinase 3 (D3). Induction of D3 by fluoxetine is inhibited by U0126 and SB203580, suggesting that Erk and p38 MAP kinases are involved. Glial-derived nerve factor (GDNF) induction by fluoxetine is prevented by U0126, suggesting that Erk is implicated. Brain-derived nerve factor (BDNF) induction seems mediated by other signaling pathways. In conclusion, we show that fluoxetine alone rapidly activates mitogen activated protein (MAP) kinase cascades in rat astrocytes and that genes involved in neuroprotection are induced in a few hours in a MAP kinase-dependent or -independent manner.

Iodotyrosine dehalogenase 1 (DEHAL1) is a transmembrane protein involved in the recycling of iodide close to the thyroglobulin iodination site.

In the thyroid, iodotyrosine dehalogenase acts on the mono and diiodotyrosines released during the hydrolysis of thyroglobulin to liberate iodide, which can then reenter the hormone-producing pathways. It has been reported that the deiodination of iodotyrosines occurs predominantly in the microsomes and is mediated by NADPH. Recently, two cDNAs, 7401- and 7513-base pairs long that encode proteins with a conserved nitroreductase domain were published in GenBank as iodotyrosine dehalogenase 1 (DEHAL1) and iodotyrosine dehalogenase 1B (DEHAL1B), respectively. We report here our investigation of the localization and activity of one of these isoforms, DEHAL1. DEHAL1 mRNA is highly expressed in the thyroid, is up-regulated by cAMP, and encodes a transmembrane protein that efficiently catalyzes the NADPH-dependent deiodination of mono (L-MIT) and diiodotyrosine (L-DIT), with greater activity vs. L-MIT. Iodotyrosine deiodinase was active in HEK293 cells transfected by DEHAL1 cDNA, but not in CHO cells. A fraction of DEHAL1 protein is exposed to the cell surface, as indicated by biotinylation experiments. Immunohistochemistry studies showed that DEHAL1 proteins accumulate at the apical pole of thyrocytes. Taken together, these findings indicate that the deiodination reaction occurs at the apical pole of the thyrocyte and is involved in a rapid iodide recycling process at and/or close to the organification site.