Protective and regenerative therapies of the CNS

Fernando Pitossi - Fundación Instituto Leloir

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The general aim of our laboratory is to provide information to design protective or regenerative therapies for Parkinson´s Disease (PD).  Unfortunately, there is no current therapy that modifies disease progression. The death of the dopaminergic neurons of the nigrostriatal system is the primary cause of the motor symptoms observed in these patients. Microglial activation in the nigrostriatal system is ubiquitously found in animal models of PD and PD patients.

Our laboratory has been focusing on the study of neuroinflammation in PD during the last 17 years to identify new therapeutic targets that could be the basis of protective therapies against this disease. The first question we wanted to ask was: is microglial activation only a consequence of neuronal death or it has functional effects on neuronal survival and therefore could be regarded as a therapeutic target? At the beginning, we have observed that microglial activation possesses toxic as well as neuroprotective effects during dopaminergic degeneration and that the cytokines secreted by them are instrumental to these effects. Therefore, our second question was: which factors determine univocal effects (toxic or neuroprotective) of brain cytokines during neurodegeneration? Searching for this answer, we have generated 5 animal models of neurodegeneration by the modulation of neuroinflammation by cytokines. In these models, the magnitude, duration and timing of cytokine expression allow to predict a univocal effect, enabling in depth studies on the mechanism of action implicated. We hypothesize that mediators exist that are common to the toxic effects of cytokines in these 5 models generated by different pro-inflammatory stimuli. We pretend to identify those common molecules downstream of the inflammatory stimuli by functional genomics in the models generated; becoming independent from the factors that alter the final functional effect of the cytokines.

On the other hand, since 2000 we study adult neural stem cells (NSC) as a putative source for regenerative therapies for PD. In particular, we study the effects of cytokines on the proliferation, differentiation and survival of NSC endogenously and after transplantation in the brain. We have observed a pro-neurogenic effect of TGF-beta 1, an anti-inflammatory cytokine, on NSC in vivo and in vitro in 3 different experimental models. Based on these and other results, the field has incorporated cytokines as part of the neurogenic niche. We are now focusing in studying the mechanism of action of TGF-beta1 in vivo and in vitro by functional genomics.

In addition, we have developed the technology of cellular reprogramming as a source of dopaminergic neurons from fibroblasts for future regenerative therapies and as in vitro models of PD. This technology will allow merging all the information and animal and cellular models that we have generated. On one hand, the therapeutic targets for protective therapies that we identify will be tested on dopaminergic neurons derived from iPS cells from PD patients. On the other, every cell transplantation in the brain will elicit inflammation. Therefore, the information on the functional effects of cytokines on cell proliferation, differentiation and survival will be investigated on dopaminergic neurons transplanted into traditional PD models or PD models based on neuroinflammation generated in our laboratory.  We are currently members of a consortium with the California Institute for Regenerative Medicine and the NIH to design and implement a clinical trial against PD using dopaminerig neurons derived from pluripotent cells.
Aim 1. Identify molecular mediators of the toxic effects of neuroinflammation in PD
As an effort to develop neuroprotective therapies against PD, we seek to identify inflammation-related molecules that could be defined as therapeutic targets. We hypothesize that there are common mediators to the toxic effects of inflammation in the nigrostriatal system. We are applying functional genomics on 5 different animal models where inflammation triggered by several inflammatory stimuli generates nigrostriatal neurodegeneration and motor symptoms (for more detailed information on these models, refer to: Neurobiology of Disease, 2006 Oct;24(1):183-93; Brain, 2008, 131:1880-94, Neurobiology of Disease, 2010, 37(3):630-40 J Neuroimmunol. 2010, May;222(1-2):29-39 and Exp Neurol. 2011 Feb;227(2):237-51). The toxic effects of the identified genes will be tested on dopaminergic neurons derived from iPS cells and animal models of PD.

Aim 2. Identify the molecular mechanism of the pro-neurogenic effect of TGF-beta1
We have identified and validated differential genes that are candidates to mediate the pro-neurogenic effects of TGF-beta1 by functional genomics. We plan to incorporate this information on the differentiation protocol of neuronal precursors derived from iPS cells already generated in our laboratory.

Aim 3. Regenerative therapies: pre-clinical studies
We seek to test the safety and efficacy of the transplantation of dopaminergic neurons derived from iPS cells on animal models of PD. We also plan to improve the safety and efficacy of the protocol by including anti-inflammatory treatments and/or expression of genes related to inflammation.

Leal MC, Casabona JC, Puntel M, Pitossi FJ.  Interleukin-1β and tumor necrosis factor-α: reliable targets for protective therapies in Parkinson's Disease? Front Cell Neurosci. 2013 Apr 29;7:53.

Graciarena M, Roca V, Mathieu P, Depino AM, Pitossi FJ. Differential vulnerability of adult neurogenesis by adult and prenatal inflammation: Role of TGF-β1. Brain Behav Immun. 2013 Nov;34:17-28

Anthony DC, Pitossi FJ. Editorial: The changing face of inflammation in the brain. Mol Cell Neurosci. 2012 Nov 10. doi:pii: S1044-7431(12)00204-7. Factor de impacto: 3,861 PubMed

Murta V, Pitossi FJ, Ferrari CC. CNS response to a second pro-inflammatory event depends on whether the primary demyelinating lesion is active or resolved. Brain Behav Immun. 2012 Oct;26(7):1102-15. PubMed

Campolongo M, Benedetti L, Podhajcer OL, Pitossi F, Depino AM. Hippocampal SPARC regulates depression-related behavior. Genes Brain Behav.; 2012; Sep 5. doi: 10.1111/j.1601-183X.2012.00848.x PubMed

Mathieu P, Roca V, Gamba C, Del Pozo A, Pitossi F. *.* autor correspondiente. Neuroprotective effects of human umbilical cord mesenchymal stromal cells in an immunocompetent animal model of Parkinson's disease. J Neuroimmunol. 2012 May 15;246(1-2):43-50. Epub 2012 Mar 27. PubMed

Leal MC, Surace EI, Holgado MP, Ferrari CC, Tarelli R, Pitossi F, Wisniewski T, Castaño EM, Morelli L. Notch signaling proteins HES-1 and Hey-1 bind to insulin degrading enzyme (IDE) proximal promoter and repress its transcription and activity: Implications for cellular Aβ metabolism. Biochim Biophys Acta, 2012 Feb;1823(2):227-35. PubMed

Barcia C, Hunot S, Guillemin GJ, Pitossi F. Editorial: Inflammation and Parkinson's disease. Parkinsons Dis. 2011;2011:729054. PubMed

Roca V, Casabona JC, Radice P, Murta V, Pitossi FJ*.* autor correspondiente. The degenerating substantia nigra as a susceptible region for gene transfer-mediated inflammation. Parkinsons Dis. 2011; 2011:931572. Review. PubMed

Depino AM, Lucchina L, Pitossi F. Early and adult hippocampal TGF-β1 overexpression have opposite effects on behavior. Brain Behav Immun. 2011, 25(8):1582-91 PubMed

Irene RE Taravini, Mariela Chertoff, Eduardo G Cafferata, José Courty, Gustavo M Murer, Fernando J Pitossi* y Oscar S Gershanik*. * contribución similar. Pleiotrophin over-expression provides trophic support to dopaminergic neurons in parkinsonian rats Molecular Neurodegeneration, 2011;6(1):40. PubMed

Clara García Samartino, M. Victoria Delpino, Clara Pott Godoy, María Silvia Di Genaro, Karina A. Pasquevich, Astrid Zwerdling, Paula Barrionuevo, Patricia Mathieu, Juliana Cassataro, Fernando J. Pitossi y Guillermo H. Giambartolomei. Brucella abortus Induces The Secretion of Pro-inflammatory Mediators From Glial Cells Leading to Astrocyte Apoptosis. American Journal of Pathology, 2010, 176(3):1323-38. PubMed

Lucchina L, Carola V, Pitossi F, Depino AM. Evaluating the interaction between early postnatal inflammation and maternal care in the programming of adult anxiety and depression related behaviors. Behav Brain Res. 2010;213(1):56-65. PubMed

Taylor PL, Barker RA, Blume KG, Cattaneo E, Colman A, Deng H, Edgar H, Fox IJ, Gerstle C, Goldstein LS, High KA, Lyall A, Parkman R, Pitossi FJ, Prentice ED, Rooke HM, Sipp DA, Srivastava A, Stayn S, Steinberg GK, Wagers AJ, Weissman IL. Patients beware: commercialized stem cell treatments on the web. Cell Stem Cell. 2010 Jul 2;7(1):43-9. Artículo publicado como resultado del trabajo del Grupo de Trabajo de la International Society for Stem Cell Research (ISSCR) sobre terapias no aprobadas utilizando células madre (turismo de células madre). Como consecuencia de esa labor, se generó una página web (www.closerlookatstemcells.org) para ayudar a la población a informarse sobre nuevos tratamientos utilizando células madre. PubMed

Garciarena, M., Depino, A., Pitossi, F.J. Prenatal inflammation impairs adult neurogenesis and memory related behavior through persistent hippocampal TGFb1 downregulation.  Brain Behav Imm. (en prensa). PubMed

De Lella Ezcurra, A.L., Chertoff, M., Graciarena, M., Ferrari, C., Pitossi, F.J. Chronic expresión of low levels of Tumor necrosis factor alpha in the substantia nigra elicits progressive neurodegeneration delayed motor symptoms and microglia/macrophage activation.  Neurobiol Dis. 37: 630-640 (2010).     PubMed

Mathieu, P., Battista, D., Depino, A., Roca, V., Pitossi, F.J. The more you have, the less you get: the functional role of inflammation on neuronal differentiation of endogenous and transplanted neural stem cells in the adult brain.  J Neurochem. 112: 1368-1385 (2010).     PubMed

Pott Godoy, M.C., Ferrari, C.C., Pitossi, F.J. Nigral neurodegeneration triggered by striatal AdlL-1 administration can be exacerbated by systemic expression.  J Neuroimmunol. 222: 29-39 (2010).     PubMed

Mathieu P, Piantanida AP, Pitossi F*. * autor correspondiente. Chronic expression of transforming growth factor-beta enhances adult neurogenesis. Neuroimmunomodulation. 2010;17(3):200-1. PubMed

Chertoff M, Di Paolo N, Schoeneberg A, Depino A, Ferrari C, Wurst W, Pfizenmaier K, Eisel U, Pitossi F*. * autor correspondiente.. Neuroprotective and neurodegenerative effects of the chronic expression of tumor necrosis factor α in the nigrostriatal dopaminergic circuit of adult mice. Exp Neurol. 2011 Feb;227(2):237-51. PubMed

Pott Godoy, M.C., Tarelli, R., Ferrari, C.C., Sarchi, M.I.S., Pitossi, F.J. Central and systemic IL-1 exacerbates neurodegeneration and motor symptoms in a model of Parkinson’s disease.  Brain. 131: 1880-1894 (2008).    PubMed

Battista, D., Ferrari, C.C., Gage, F.H., Pitossi, F.J. Neurogenic niche modulation by activated microglia: transforming growth factor beta increases neurogenesis in the adult dentate gyrus. Eur. J. Neurosci. 23: 83-93 (2006).     PubMed

Ferrari, C.C., Depino, A.M., Prada, F., Muraro, N., Campbell, S., Podhajcer, O.L., Perry, V.H., Anthony, D.C.,Pitossi, F.J. Reversible demyelination, blood-brain barrier breakdown, and pronounced neutrophil recruitment induced by chronic IL-1 expression in the brain. Am. J. Pathol. 165: 1827-1837 (2004).     PubMed

M. Kolb, P. J. Margetts, D. C. Anthony, F. Pitossi y J. Gauldie, Transient expression of IL-1 beta induces acute lung injury and chronic repair leading to pulmonary fibrosis.(2001). J. of Clinical Investigation, 107, 12, p. 1529- 1536. PubMed

H. Schneider, F. Pitossi, D. Balschun, A. Wagner, A. del Rey, y H.O. Besedovsky ,neuromodulatory role of interleukin-1 beta in the hippocampus. (1998) Proceedings of the National Academy of Sciences of the United States of America, 95, 7778-7783. PubMed

Pitossi, F.J., Blank, A., Schröder, A., Schwarz, A., Hüssi, P. Schwemmle, M., Pavlovic, J., Staeheli, P. A functional GTP-binding motif is necessary for antiviral activity of Mx proteins.  J Virol. 67: 6726-6732 (1993).     PubMed

P. Jones, T. Jakubowicz, F. Pitossi, F. Maurer y B. Hemmings. Molecular cloning and identification of a serine-threonine protein kinase of the 2nd. messanger subfamily. Proceedings of the National Academy of Sciences of the United States of America (1991). Vol 88, No 10, págs.: 4171-4175. PubMed





Fernando Pitossi
Head of Laboratory - fpitossi@leloir.org.ar



María Celeste Leal
Assistant Research - CONICET



Berenice Silva
PhD Student - Fundación René Baron



Brenda Erhardt
Ph.D. Student - CONICET



Esteban Miglietta
Ph.D. Student - CONICET



María Isabel Farias
Technical assistant



Victoria Gradaschi
Passant