Ispitivanje ranih markera procesa neuroinflamacije i oksidativnog stresa u mozgu transgenog hSOD1 G93A pacova kao eksperimentalnog modela amiotrofične lateralne skleroze

Abstract

Amiotrofična lateralna skleroza je smrtonosno neurodegenerativno oboljenje koje pogađa motorne i kognitivne domene centralnog nervnog sistema (CNS). Mutacije u genu za Cu, Zn-supeoksid dismutazu (SOD1) uzrokuju 20% familijalnih slučajeva ALS i izazivaju stvaranje unutarćelijskih SOD1 agregata i smanjeno vezivanje Cu i Zn, što dovodi do patoloških unutarćelijskih promena i neurodegeneracije. Istraživanja na post mortem tkivu ALS pacijenata i životinjskim modelima bolesti su istakla glijalnu aktivaciju, poremećenu homeostazu prelaznih metala, oštećenje krvno-moždane barijere (KMB) i povećanu proizvodnju reaktivnih kiseoničnih vrsta kao glavne patološke faktore u bolesti. Cilj ove studije je bio da se ispitaju navedene patološke promene u motornim (moždano stablo) i ne-motornim (hipokampus) centrima u mozgu transgenog hSOD1 G93A pacovskog modela ALS, u presimptomatskom i simptomatskom stadijumu bolesti, kako bi se detaljnije proučili ovi patofiziološki mehanizmi i identifikovale rane promene u bolesti. Prisustvo i vremenski sled aktivacije i morfoloških promena mikroglije i astrocita, kao i akumulacija i distribucija SOD1 proteina u ovim ćelijama su ispitani naprednim 3D modelovanjem i analizom kolokalizacije na slikama dobijenim konfokalnom laserskom skenirajućom mikroskopijom. Rezultati su pokazali glijalnu proliferaciju i progresivno nakupljanje SOD1 u tkivu u oba ispitivana moždana regiona hSOD1 G93A pacova, koji počinju već u presimptomatskoj fazi. U moždanom stablu ovih životinja aktivacija astrocita je bila prisutna pre početka simptoma bolesti, i bila je praćena aktivacijom mikroglije. Takođe, u moždanom stablu astrociti su pokazali progresivnu akumulaciju SOD1 proteina u ćelijskom telu i nastavcima, dok su nivoi SOD1 u mikrogliji bili smanjeni a njena distribucija ograničena na distalne ćelijske nastavke. U hipokampusu hSOD1 G93A pacova astrociti su pokazali isti reaktivni profil kao onaj uočen u moždanom stablu, dok je morfologija mikroglije bila nepromenjena. U ovom regionu mozga oba tipa glijalnih ćelija su pokazala akumulaciju SOD1 proteina u ćelijskom telu...

Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disorder affecting both the motor and cognitive domains of the central nervous system (CNS). Mutations in the Cu,Zn-superoxide dismutase (SOD1) cause 20% of familial ALS and provoke formation of intracellular SOD1 aggregates and copper and zinc unbinding, leading to pathological intracellular changes and neurodegeneration. Post mortem and animal model studies indicate glial activation, perturbed transition metal homeostasis, blood-brain barrier (BBB) disruption and elevated production of reactive oxygen species as major contributors to disease pathology. The aim of this study was to investigate the above mentioned pathological changes in motor (brainstem) and non-motor (hippocampus) brain centers of a transgenic hSOD1 G93A rat model of ALS, in presymptomatic and symptomatic stages of the disease, in order to further clarify these pathophysiological mechanisms and to identify early changes in the disease. The presence and the timeline of activation and morphological changes of microglia and astrocytes, as well as their intracellular accumulation and distribution of SOD1 were investigated by advanced 3D modeling and colocalization analysis of images obtained by confocal laser scanning microscopy. Results revealed glial proliferation and progressive tissue accumulation of SOD1 in both brain regions of hSOD1 G93A rats starting already at the presymptomatic stage. In the brainstem of these animals astrocytes activation occurred before disease symptoms onset, and was followed by activation of microglia. Additionally, in the brainstem astrocytes demonstrated progressive SOD1 accumulation in the cell body and processes, while microglial SOD1 levels were reduced and its distribution limited to distal cell processes. In the hippocampus of hSOD1 G93A rats astrocytes exhibited an identical reactive profile as the one observed in the brainstem, while microglial morphology was unchanged. In this brain region both glial cell types exhibited SOD1 accumulation in the cell body...