Human central nervous system astrocytes support survival and activation of B cells: implications for MS pathogenesis.

BACKGROUND: The success of clinical trials of selective B cell depletion in patients with relapsing multiple sclerosis (MS) indicates B cells are important contributors to peripheral immune responses involved in the development of new relapses. Such B cell contribution to peripheral inflammation likely involves antibody-independent mechanisms. Of growing interest is the potential that B cells, within the MS central nervous system (CNS), may also contribute to the propagation of CNS-compartmentalized inflammation in progressive (non-relapsing) disease. B cells are known to persist in the inflamed MS CNS and are more recently described as concentrated in meningeal immune-cell aggregates, adjacent to the subpial cortical injury which has been associated with progressive disease. How B cells are fostered within the MS CNS and how they may contribute locally to the propagation of CNS-compartmentalized inflammation remain to be elucidated. METHODS: We considered whether activated human astrocytes might contribute to B cell survival and function through soluble factors. B cells from healthy controls (HC) and untreated MS patients were exposed to primary human astrocytes that were either maintained under basal culture conditions (non-activated) or pre-activated with standard inflammatory signals. B cell exposure to astrocytes included direct co-culture, co-culture in transwells, or exposure to astrocyte-conditioned medium. Following the different exposures, B cell survival and expression of T cell co-stimulatory molecules were assessed by flow cytometry, as was the ability of differentially exposed B cells to induce activation of allogeneic T cells. RESULTS: Secreted factors from both non-activated and activated human astrocytes robustly supported human B cell survival. Soluble products of pre-activated astrocytes also induced B cell upregulation of antigen-presenting cell machinery, and these B cells, in turn, were more efficient activators of T cells. Astrocyte-soluble factors could support survival and activation of B cell subsets implicated in MS, including memory B cells from patients with both relapsing and progressive forms of disease. CONCLUSIONS: Our findings point to a potential mechanism whereby activated astrocytes in the inflamed MS CNS not only promote a B cell fostering environment, but also actively support the ability of B cells to contribute to the propagation of CNS-compartmentalized inflammation, now thought to play key roles in progressive disease.

Anti-Mullerian-hormone-dependent regulation of the brain serine-protease inhibitor neuroserpin.

The balance between tissue-type plasminogen activator (tPA) and one of its inhibitors, neuroserpin, has crucial roles in the central nervous system, including the control of neuronal migration, neuronal plasticity and neuronal death. In the present study, we demonstrate that the activation of the transforming growth factor-beta (TGFbeta)-related BMPR-IB (also known as BMPR1B and Alk6)- and Smad5-dependent signalling pathways controls neuroserpin transcription. Accordingly, we demonstrate for the first time that anti-Mullerian hormone (AMH), a member of the TGFbeta family, promotes the expression of neuroserpin in cultured neurons but not in astrocytes. The relevance of these findings is confirmed by the presence of both AMH and AMH type-II receptor (AMHR-II) in brain tissues, and is supported by the observation of reduced levels of neuroserpin in the brain of AMHR-II-deficient mice. Interestingly, as previously demonstrated for neuroserpin, AMH protects neurons against N-methyl-D-aspartate (NMDA)-mediated excitotoxicity both in vitro and in vivo. This study demonstrates the existence of an AMH-dependent signalling pathway in the brain leading to an overexpression of the serine-protease inhibitor, neuroserpin, and neuronal survival.

Tissue-type plasminogen activator rescues neurones from serum deprivation-induced apoptosis through a mechanism independent of its proteolytic activity.

Although the mechanism of action of tissue-type plasminogen activator (tPA) in excitotoxic necrosis is well documented, whether this serine protease can influence the apoptotic cascade remains a subject of debate. Here, we report that tPA protects cultured cortical neurones against apoptotic cell death induced by serum deprivation, an effect associated with a reduction of caspase-3 activation. Interestingly, blocking tPA proteolytic activity by either tPA stop or neuroserpin did not prevent this neuroprotection. Similarly, prevention of the interaction between tPA and its receptor low-density lipoprotein receptor-related protein (LRP) could not alter tPA anti-apoptotic activity. Interestingly, the survival-promoting effect of tPA was blocked by the phosphatidylinositol-3 (PI-3) kinase inhibitor, LY294002, but not by the mitogen-activated protein (MAP) kinase inhibitor, U0126. In conclusion, the present demonstration of an anti-apoptotic effect of tPA, independent of its enzymatic activity, reveals an additional level of complexity in our understanding of this critical mediator of brain physiology and pathology.

The brain-specific tissue-type plasminogen activator inhibitor, neuroserpin, protects neurons against excitotoxicity both in vitro and in vivo.

Considering its brain-specific expression, neuroserpin (NS), a potent inhibitor of tissue-type plasminogen activator (tPA), might be a good therapeutic target to limit the pro-excitotoxic effects of tPA within the cerebral parenchyma, without affecting the benefit from thrombolysis in stroke patients. Here, we aimed at determining the mechanisms of action responsible for the previously reported neuroprotective activity of NS in rodent experimental cerebral ischemia. First, we show in vivo that exogenous NS protects the cortex and the striatum against NMDA-induced injury. Then, the cellular mechanisms of this neuroprotection were investigated in primary cultures of cortical neurons. We show that NS fails to prevent serum deprivation-induced apoptotic neuronal death, while it selectively prevents NMDA- but not AMPA-induced excitotoxicity. This beneficial effect is associated to a decrease in NMDA receptor-mediated intracellular calcium influx. Altogether, these data suggest that an overexpression of neuroserpin in the brain parenchyma might limit the deleterious effect of tPA on NMDA receptor-mediated neuronal death, which occurs following experimental ischemia.

Cytokines in neuroinflammation and Alzheimer's disease.

Inflammation has been reported in numerous neurodegenerative disorders such as Parkinson's disease, stroke and Alzheimer's disease (AD). In AD, the inflammatory response is mainly located to the vicinity of amyloid plaques. Cytokines, such as Interleukin-1 (IL-1), Interleukin-6 (IL-6), Tumor Necrosis Factor alpha (TNF-alpha) and Transforminng Growth Factor beta (TGF-beta) have been clearly involved in this inflammatory process. Although their expression is induced by the presence of amyloid-beta peptide, these cytokines are also able to promote the accumulation of amyloid-beta peptide. Altogether, IL-1, IL-6, TNF-alpha and TGF-beta should be considered as key players of a vicious circle leading to the progression of the disease.

Sp1 and Smad transcription factors co-operate to mediate TGF-beta-dependent activation of amyloid-beta precursor protein gene transcription.

Abnormal deposition of Abeta (amyloid-beta peptide) is one of the hallmarks of AD (Alzheimer's disease). This peptide results from the processing and cleavage of its precursor protein, APP (amyloid-beta precursor protein). We have demonstrated previously that TGF-beta (transforming growth factor-beta), which is overexpressed in AD patients, is capable of enhancing the synthesis of APP by astrocytes by a transcriptional mechanism leading to the accumulation of Abeta. In the present study, we aimed at further characterization of the molecular mechanisms sustaining this TGF-beta-dependent transcriptional activity. We report the following findings: first, TGF-beta is capable of inducing the transcriptional activity of a reporter gene construct corresponding to the +54/+74 region of the APP promoter, named APP(TRE) (APP TGF-beta-responsive element); secondly, although this effect is mediated by a transduction pathway involving Smad3 (signalling mother against decapentaplegic peptide 3) and Smad4, Smad2 or other Smads failed to induce the activity of APP(TRE). We also observed that the APP(TRE) sequence not only responds to the Smad3 transcription factor, but also the Sp1 (signal protein 1) transcription factor co-operates with Smads to potentiate the TGF-beta-dependent activation of APP. TGF-beta signalling induces the formation of nuclear complexes composed of Sp1, Smad3 and Smad4. Overall, the present study gives new insights for a better understanding of the fine molecular mechanisms occurring at the transcriptional level and regulating TGF-beta-dependent transcription. In the context of AD, our results provide additional evidence for a key role for TGF-beta in the regulation of Abeta production.

The complexity of tissue-type plasminogen activator: can serine protease inhibitors help in stroke management?

Stroke, the third leading cause of death in industrialised countries, represents a major burden on healthcare authorities. The elucidation of molecular events sustaining infarct evolution in experimental models has allowed the development of putative therapeutic agents. However, despite marked benefits in animals, most of them have failed in clinical trials. At present, the only approved therapy for stroke is early reperfusion by intravenous injection of the thrombolytic agent, tissue-type plasminogen activator (tPA). tPA-dependent thrombolysis sometimes promotes haemorrhage, but improves neurological outcome in a great proportion of patients, provided it is performed within the recommended therapeutic window. In addition to the benefit of tPA injection in the vascular compartment, this endogenously produced serine protease could also promote excitotoxic processes within the cerebral parenchyma. This article reviews the various aspects of tPA during stroke, and discusses potential improvements to current clinical management, with a particular emphasis on targeting the deleterious actions of tPA through endogenous serine protease inhibitors (serpins).