Nivo izražanja enolaze pri zorenju in vnetni stimulaciji oligodentrocitov HOG

Jenko, Manca (author), Pišlar, Anja (mentor), Horvat, Selena (comentor)

Nevrodegenerativne bolezni predstavljajo pomemben svetovni zdravstveni problem, vendar še nimamo učinkovitih terapij, ki bi upočasnile, zavrle ali preprečile katerokoli od teh bolezni, zato je pridobivanje informacij o molekularnih mehanizmih povezanih s patologijo nevrodegenerativnih bolezni bistvenega pomena. Pri nastanku in razvoju številnih nevrodegenerativnih bolezni imajo pomemebno vlogo tudi oligodendrociti. V magistrski nalogi smo proučevali nivo izražanja izooblik enolaze v oligodendrocitih, saj izooblika γ-enolaze izkazuje nevrotrofičnim dejavnikom podobno delovanje, vendar njena vloga v oligodendrocitih še ni povsem znana. V ta namen smo izbrali celično linijo oligodendrocitov humanega oligodendroglioma (HOG) in z uporabo različnih sestav diferenciacijskih gojišč spodbudili njihovo diferenciacijo, kar se je videlo v celični morfologiji, upadu celične proliferacije in spremembah nivoja izražanja dejavnikov nezrelih in zrelih oligodendrocitov. Pokazali smo, da med diferenciacijo oligodendrocitov pride do α-γ enolaznega preklopa, pri čemer se je povišal nivo celokupne kot tudi aktivne oblike γ-enolaze in znatno zmanjšal nivo α-enolaze, hkrati je razviden tudi upad glikolitične aktivnosti enolaze. Nadalje smo ugotovili, da ima zaviranje katepsina X s spojino AMS36 dodaten vpliv na diferenciacijo celic, saj je spodbudilo rast izrastkov v dolžino in preoblikovanje celic v podolgovat fenotip, kar nakazuje na posredno nevrotrofično vlogo γ-enolaze, ki je substrat katepsina X v možganskih celicah, kar podpira tudi pokazana ko-lokalizacija katepsina X in γ-enolaze v oligodendrocitih HOG. Nato smo s citokinoma dejavnikom tumorske nekroze α (TNF-α) in interferonom γ (IFN-γ) postavili vnetni model poškodovanih oligodendrocitov. Ugotovili smo, da spodbujeno vnetje povzroča smrt celic in upad nivoja izražanja α-enolaze in γ-enolaze v oligodendrocitih ter posledično zaviranje celične proliferacije in diferenciacije. V okviru magistrske naloge smo torej pokazali prisotnost α-γ enolaznega preklopa tekom diferenciacije oligodendrocitov, pri čemer se v diferenciranih celicah HOG poveča nivo izražanja γ-enolaze, za katero je značilno izkazovanje trofične podpore in je uravnavana s proteolitičnim delovanjem katepsina X. Zaradi slednjega bi razvoj spojin, ki bi s spodbujanjem trofične aktivnosti γ-enolaze spodbujale aktivacijo oligodendrocitnih predniških celic in diferenciacijo ter obnovo oligodendrocitov, predstavljal pomemben pristop za preprečevanje in zdravljenje nevrodegenerativnih bolezni.

• Language: Slovenian
• Keywords: oligodendrociti, diferenciacija, enolaza, katepsin X, nevrotrofična aktivnost
• Work type: Master's thesis/paper
• Organization: FFA - Faculty of Pharmacy
• Year: 2024
• PID: 20.500.12556/RUL-158516
• Publication date in RUL: 14.06.2024

Secondary language

• Language: English
• Title: Expression of enolase during maturation and inflammatory stimulation of HOG oligodentrocytes

Abstract

Neurodegenerative diseases pose an important global health challenge but currently there are no efficient therapies that would slow, stop or prevent these diseases, making it essential to understand molecular mechanisms related to pathology of neurodegenerative diseases. Oligodendrocytes play a crucial role in development and progression of numerous neurodegenerative diseases. In our work, we studied the expression of γ-enolase in oligodendrocytes, which exerts neurotrophic-like properties and its role has been relatively unexplored in oligodendrocytes. For this purpose, we used human oligodendroglioma (HOG) cell line, stimulating differentiation using various differentiation media compositions, with changes shown in cell morphology, decreased cell proliferation and altered expression levels of markers for undifferentiated and differentiated oligodendrocytes. We observed an α-γ enolase switch during oligodendrocyte differentiation, with an increase in the total and active forms of γ-enolase and a decrease in α-enolase expression, coinciding with decreased glycolytic activity. Inhibition of cathepsin X with AMS36 further affected cell differentiation, leading to enhanced growth of cell extensions and cell reorganizing into an elongated phenotype. This indicates indirect neurotrophic activity of cathepsin X's substrate γ-enolase in brain cells, further supported by the colocalization of cathepsin X and γ-enolase in HOG oligodendrocytes. We also established an inflammatory model of damaged oligodendrocytes using the cytokines tumour necrosis factor α (TNF-α) and interferon γ (IFN-γ). We confirmed that inflammation leads to cell death and showed a decline in α-enolase and γ-enolase expression in oligodendrocytes, resulting in inhibited cell proliferation and differentiation. The findings of this master's thesis show the presence of an α-γ enolase switch during oligodendrocyte differentiation, with increased expression levels of γ-enolase in differentiated oligodendrocytes, the isoenzyme known for its trophic support that is regulated by proteolytic activity of cathepsin X. This suggests a potential for the development of new drugs that would promote the neurotrophic activity of γ-enolase, thereby promoting the activation of oligodendrocyte progenitor cells and oligodendrocyte differentiation and renewal, which presents a promising approach for prevention and therapy of neurodegenerative diseases.

• Keywords: oligodendrocytes, differentiation, enolase, cathepsin X, neurotrophic activity