Ispitivanje terapijskog dejstva arilpiperazinskih dopaminergičkih liganada u modelu multiple skleroze kod pacova

Abstract

U ovom radu je ispitivano dejstvo dva novosintetisana arilpiperazinska D2/5-HT1A dopaminergička/serotoninergička liganda, koji ispoljavaju neuroprotektivna svojstva in vitro, u eksperimentalnom autoimunskom encefalomijelitisu (EAE) kao modelu multiple skleroze kod pacova. Oba jedinjenja, N-{4-[2-(4-phenyl-piperazin-1-yl)- ethyl}-phenyl]-picolinamide (6a) i N-{3-[2-(4-phenyl-piperazin-1-yl)-ethyl]-phenyl}- picolinamide (6b), su u dozi od 10 mg/kg i.p. ispoljila pozitivan efekat na klinički tok EAE-a kod Dark Agouti pacova imunizovanih homogenatom kičmene moždine (KM), pri čemu je derivat 6b ispoljavao nešto izraženiji efekat u odlaganju početka bolesti i snižavanju maksimalne kliničke težine, što je u skladu sa njegovim višim afinitetom za D2 i 5-HT1A receptore. Povoljan efekat na klinički tok EAE-a se zadržavao ukoliko je tretman vršen tokom efektorske faze bolesti (od 8. dana pa na dalje), ali ne i tokom induktivne faze (od 0. do 7. dana) EAE-a. Arilpiperazini su smanjivali infiltraciju CNSa inflamatornim ćelijama, kao i ekspresiju iRNK za proinflamatorne citokine TNF, IL-6, IL-1, i GM-CSF, TH1 citokin IFN-γ, TH17 citokin IL-17, kao i za glavne transkripcione faktore TH1 (T-bet) i TH17 (RORγt) ćelija u tkivu CNS-a ali nisu uticali na ekspresiju proinflamatornih medijatora u infiltratu mononuklearnih ćelija, niti su menjali njegov ćelijski sastav. Tretman arilpiperazinima je smanjio apoptotsku ćelijsku smrt i povećao aktivnost kinaze Akt (engl. v-Akt Murine Thymoma Viral Oncogene), važne u preživljavanju ćelija, i supstrata glavnog inhibitora autofagije mTOR-a (engl. mammalian target of rapamycin), p70S6K kinaze (engl. ribosomal protein S6 kinase) u CNS-u životinja obolelih od EAE-a. Konačno, u in vitro kokultivaciji oligodendrocita i neurona sa stimulisanim T limfocitima, arilpiperazini su ispoljili neuroprotektivan efekat prema oligodendrocitnoj (OLN-93) i neuronskoj (PC12) ćelijskoj liniji, spasavajući ih od normalnih T ćelija aktiviranih mitogenom i encefalitogenih T ćelija stimulisanih mijelin-baznim proteinom, ali nisu uticali na imunomodulatornu aktivnost T limfocita...

In the present study we investigated the effect of the two newly synthesized arylpiperazine D2/5-HT1A dopaminergic/serotoninergic ligands, which display neuroprotective properties in vitro, in experimental autoimmune encephalomyelitis (EAE), as an animal model of multiple sclerosis. Both compounds, N-{4-[2-(4-phenylpiperazin- 1-yl)-ethyl}-phenyl]-picolinamide (6a) and N-{3-[2-(4-phenyl-piperazin-1- yl)-ethyl]-phenyl}-picolinamide (6b), at 10 mg/kg i.p, reduced clinical signs of EAE in the spinal cord homogenate-immunized Dark Agouti rats. Compound 6b had somewhat pronounced effect in delaying the disease onset and reducing the maximal clinical score, which correlated with its higher affinity for D2 and 5-HT1A receptors. The protection was retained if the treatment was limited to the effector (from day 8 onwards), but not the inductive (day 0-7) phase of EAE. Arylpiperazines reduced CNS immune cell infiltration and expression of mRNA encoding proinflammatory cytokines TNF, IL-6, IL-1, and GM-CSF, TH1 cytokine IFN-γ, TH17 cytokine IL-17, as well as the signature transcription factors of TH1 (T-bet) and TH17 (RORγt) cells in the CNS tissue, arylpiperazines did not affect the expression of mRNA for proinflammatory mediators in mononuclear cell infiltrate, neither they changed its cellular composition. Arylpiperazine treatment reduced apoptotic cell death and increased activation of the prosurvival kinase Akt (v-Akt Murine Thymoma Viral Oncogene) and p70S6K kinase (ribosomal protein S6 kinase), the substrate of the major autophagy inhibitor mTOR (mammalian target of rapamycin), in the CNS of EAE animals. Finally, the in vitro treatment with arylpiperazines protected oligodendrocyte cell line OLN-93 and neuronal cell line PC12 from mitogen-activated normal T cells or myelin basic protein-activated encephalitogenic T cells, without exerting immunomodulatory activity...