Rol de los receptores muscarínicos en la tumorigénesis mamaria
The non neuronal cholinergic system (nNCS) is constituted by acetylcholine (ACh), choline acetyl transferase and\nacetylcholinesterase, the enzymes that synthesize and degrade ACh, and nicotinic and muscarinic receptors\n(mAChR), all of them expressed in cells with non nervous origin. The nNCS has b...
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| Formato: | Tesis de maestría acceptedVersion |
| Lenguaje: | Español |
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Facultad de Farmacia y Bioquímica
2015
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| Acceso en línea: | http://repositoriouba.sisbi.uba.ar/gsdl/cgi-bin/library.cgi?a=d&c=afamaster&cl=CL1&d=HWA_1125 http://repositoriouba.sisbi.uba.ar/gsdl/collect/afamaster/index/assoc/HWA_1125.dir/1125.PDF |
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| Sumario: | The non neuronal cholinergic system (nNCS) is constituted by acetylcholine (ACh), choline acetyl transferase and\nacetylcholinesterase, the enzymes that synthesize and degrade ACh, and nicotinic and muscarinic receptors\n(mAChR), all of them expressed in cells with non nervous origin. The nNCS has been implicated in the regulation\nof cellular physiological and pathological functions. Regarding the latter, previous works reported the\nparticipation of nNCS in tumor progression. Particularly, mAChR are over-expressed in a great number of\ntumors. We demonstrated the expression of these receptors in MCF-7 cells, derived from a human mammary\nadenocarcinoma, as well as in murine mammary tumor, cells and their absence in the non tumorigenic cell line,\nMCF-10A. Furthermore, we observed that the activation of mAChR stimulates cell proliferation and migration in\nvitro and the angiogenic response in vivo.\nTaking into account these previous results, we set out to investigate whether the expression of the subtypes 3\nand 4 of mAChR in MCF-10A cells promotes the malignization of these cells. For this aim, we generated three\ncell lines by transfection, which expressed the subtypes 3 and 4 of mAChR denominated: MCF-10A-M3, MCF-\n10A-M4 and MCF-10A-M3M4. In these three cell lines, we studied the effect of cholinergic activation on cell\ncycle, and cell migration in vitro, as well as neovascularization and tumor growth in vivo. We used the MCF-7 cell\nline as positive control. We demonstrated that all cell lines transfected express mRNA and proteins for the\nsubtypes 3 and/or 4 of mAChR. We observed that the cholinergic agonist carbachol increased the percentage of\ncells in S/G2/M phase in MCF-10A-M3, MCF-10A-M4 and MCF-10A-M3M4 cells, and that this effect was reduced\nby atropine, a muscarinic non-selective antagonist.\nWe also studied the effect of the agonist on cell migration and metalloproteinase-9 (MMP-9) expression, as\nfundamental parameters in tumor invasion. The treatment with carbachol increased migration and MMP-9\nexpression in all transfected cell lines, in a similar manner to that observed in MCF-7 cells, while it had no effect\nover these parameters in MCF-10A cells. The action of carbachol was reduced by atropine and also by 4-DAMP\nor tropicamide, selective antagonists for M3 and M4 receptors, respectively.\nAngiogenesis is a central step for tumor progression and for this reason, we studied the ability of MCF-10A-M3,\nMCF-10A-M4 and MCF-10A-M3M4 cells to generate new blood vessels in vivo. We found that cells increased\nneovascularization similarly to MCF-7 cells. Also, the transfected cell lines expressed de novo vascular\nendothelial growth factor-A (VEGF-A), a constitutive property in MCF-7 cells.\nFinally we evaluated tumorigenic ability in vivo and we found that MCF-10A-M3, MCF-10A-M4 and MCF-10AM3M4\ncells generate tumors in NUDE mice, while MCF-10A cells lack of this ability.\nIn conclusion, the expression of one or more subtypes of mAChR in mammary human non-tumorigenic cells is\nenough to increase cell proliferation, migration and MMP-9 production in vitro, and to acquire the ability to\ninduce neovascularization, to produce VEGF and to generate tumors in vivo. |
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