Multiple sclerosis (MS) is an autoimmune demyelinating disease of the central nervous system (CNS), characterized by acute focal inflammatory demyelination and axonal damage induced by myelin-reactive T cells. However, not only T cells, but also other cell types of the immune system, are involved in the onset and progress of the disease. In fact, it has been proposed a role for Natural Killer (NK) cells in the relapsing-remitting progression of MS. NK cells play an essential role in controlling tumor transformation and viral infection by killing aberrant cells that down-modulates human leukocyte antigen (HLA) class I surface expression, while sparing cells with HLA-I via recognition by killer-cell inhibitory receptors (KIR) that finely tune NK cell effector functions. Epidemiological studies have shown that the genetic predisposition and the exposure to an infectious agent might be involved in MS pathogenesis. In particular, several data suggest the involvement of herpes viruses’ infection. In previous studies we have evaluated the role of NK cells towards herpes viruses (HHVs) infection in MS patients and we have observed an interesting correlation between a defective activation of NK cells towards infection and an up-modulation of KIR2DL2 inhibitory receptor expression. The aim of this project was to investigate the molecular mechanisms that differentiate NK cell behavior towards HHVs infection in association with KIR2DL2 expression and the possible implication n MS follow-up. We have analyzed the expression and effect on NK cell activation of KIR2DL2 receptor during HSV-1, HHV-6A, -6B and EBV infection. We have observed an increased activation of NK without KIR2DL2 expression in the presence of HHVs infected cells, whereas KIR2DL2+ NK cells were not able to activate. On the basis of these data, we have investigated the possible molecular and epigenetic modifications involved in the expression of KIR2DL2 receptor during HHVs infections. In particular, we have identified a different cytokine expression profile betweenKIR2DL2 positive and negative NK cells, reflecting a lower activation in KIR2DL2+ NK cells. Moreover, we have investigated a possible implication of transcription factors expression in modulating KIR2DL2 expression. We have analyzed Sp1 mRNA and protein expression in KIR2DL2 positive and negative NK cells cells and evaluated KIR2DL2 promoter interaction with transcription factor RUNX1. We have found a significant role for Sp1 in regulating and for RUNX1 in up-modulating KIR2DL2 expression. After that, we have investigated the role of epigenetic modification, methylation, in modulating the expression of KIR2DL2 in MS patients. We have observed a correlation between KIR2DL2 promoter methylation, leading to KIR2DL2 gene silencing, and a better MS prognosis. Altogether these data provide a novel point of view in the landscape of the immune response toward HHVs infection in MS disease, in particular it will be possible to exploit KIR2DL2 receptor expression as a target for modulating NK cell activation.
La sclerosi multipla (SM) è una malattia demielinizzante autoimmune del sistema nervoso centrale (CNS), caratterizzata da demielinizzazione infiammatoria acuta e danno assonale indotti da cellule T reattive alla mielina. Tuttavia, non solo le cellule T, ma anche altri fattori, appartenenti al sistema immunitario, sono coinvolti nell'insorgenza e progressione della malattia. Infatti è stato osservato un ruolo significativo delle cellule Natural Killer (NK) nelle forme recidivanti-remittenti della SM. Le cellule NK giocano un ruolo essenziale nel controllo della trasformazione tumorale così come delle infezioni virali, poiché sono in grado di uccidere le cellule aberranti che presentano una diminuita espressione dell'antigene leucocitario umano (HLA) di classe I, mentre risparmiano le cellule esprimenti HLA-I, il cui riconoscimento avviene da parte dei recettori inibitori KIR, regolatori delle funzioni effettrici delle cellule NK. Studi epidemiologici hanno dimostrato che la predisposizione genetica e l'esposizione ad un agente infettivo risultano essere coinvolti nella patogenesi della SM. In particolare, molti dati suggeriscono il coinvolgimento di infezioni da herpes virus. In studi precedenti, valutando il ruolo delle cellule NK in risposta a infezioni da herpes virus in pazienti affetti da SM, abbiamo riscontrato una correlazione interessante tra un'attivazione difettiva delle cellule NK nei confronti dell'infezione e una sovra-espressione del recettore inibitorio KIR2DL2. Lo scopo di questo progetto è stato quello di studiare i meccanismi molecolari che differenziano il comportamento delle cellule NK in risposta ad infezioni da herpes virus umani (HHVs) in associazione con l’espressione del recettore KIR2DL2. Per questo motivo, abbiamo analizzato l'espressione e la funzione di KIR2DL2 durante l’infezione da HSV-1, HHV-6A, -6B ed EBV. Abbiamo potuto riscontrare che l'attivazione delle cellule NK risultava incrementata nelle cellule NK KIR2DL2- in co-coltura con cellule infettate con HHVs, mentre le cellule NK KIR2DL2+ non risultavano attivate. Sulla base di questi dati abbiamo studiato le possibili modificazioni molecolari ed epigenetiche coinvolte. In particolare abbiamo identificato variazioni nell’espressione di differenti citochine in cellule NK KIR2DL2+ e KIR2DL2-, che riflettono una minore risposta immunitaria in cellule NK esprimenti il recettore KIR2DL2. Inoltre abbiamo indagato una possibile implicazione dei fattori di trascrizione nel modulare l'espressione del KIR2DL2, analizzando l’mRNA e la proteina di Sp1 in cellule KIR2DL2 positive e negative e valutando l'interazione del promotore del recettore KIR2DL2 con il fattore di trascrizione RUNX1. Si è potuto riscontrare un ruolo significativo di entrambi nell’espressione del KIR2DL2, in particolare di Sp1 nella regolazione e di RUNX1 nella sovra-stimolazione dell’espressione del recettore. In seguito ci siamo occupati di investigare il ruolo delle modificazioni epigenetiche, in particolare della metilazione, nel modulare l'espressione del KIR2DL2 in pazienti con SM. Sulla base dei risultati ottenuti abbiamo ipotizzato che La metilazione del promotore del KIR2DL2 in pazienti con SM, che porta al silenziamento del gene, sembra correlare con una migliore prognosi di malattia. Complessivamente questi dati forniscono un nuovo punto di vista nello studio della risposta immunitaria verso infezioni da herpes virus in pazienti affetti da SM. In particolare, sarà possibile sfruttare l'espressione del recettore KIR2DL2 come target per la modulazione dell'attivazione delle cellule NK.
NK cells expressing KIR2DL2 inhibitory receptor: effect on HHVs infection control in MS disease
BOLZANI, Silvia
2017
Abstract
Multiple sclerosis (MS) is an autoimmune demyelinating disease of the central nervous system (CNS), characterized by acute focal inflammatory demyelination and axonal damage induced by myelin-reactive T cells. However, not only T cells, but also other cell types of the immune system, are involved in the onset and progress of the disease. In fact, it has been proposed a role for Natural Killer (NK) cells in the relapsing-remitting progression of MS. NK cells play an essential role in controlling tumor transformation and viral infection by killing aberrant cells that down-modulates human leukocyte antigen (HLA) class I surface expression, while sparing cells with HLA-I via recognition by killer-cell inhibitory receptors (KIR) that finely tune NK cell effector functions. Epidemiological studies have shown that the genetic predisposition and the exposure to an infectious agent might be involved in MS pathogenesis. In particular, several data suggest the involvement of herpes viruses’ infection. In previous studies we have evaluated the role of NK cells towards herpes viruses (HHVs) infection in MS patients and we have observed an interesting correlation between a defective activation of NK cells towards infection and an up-modulation of KIR2DL2 inhibitory receptor expression. The aim of this project was to investigate the molecular mechanisms that differentiate NK cell behavior towards HHVs infection in association with KIR2DL2 expression and the possible implication n MS follow-up. We have analyzed the expression and effect on NK cell activation of KIR2DL2 receptor during HSV-1, HHV-6A, -6B and EBV infection. We have observed an increased activation of NK without KIR2DL2 expression in the presence of HHVs infected cells, whereas KIR2DL2+ NK cells were not able to activate. On the basis of these data, we have investigated the possible molecular and epigenetic modifications involved in the expression of KIR2DL2 receptor during HHVs infections. In particular, we have identified a different cytokine expression profile betweenKIR2DL2 positive and negative NK cells, reflecting a lower activation in KIR2DL2+ NK cells. Moreover, we have investigated a possible implication of transcription factors expression in modulating KIR2DL2 expression. We have analyzed Sp1 mRNA and protein expression in KIR2DL2 positive and negative NK cells cells and evaluated KIR2DL2 promoter interaction with transcription factor RUNX1. We have found a significant role for Sp1 in regulating and for RUNX1 in up-modulating KIR2DL2 expression. After that, we have investigated the role of epigenetic modification, methylation, in modulating the expression of KIR2DL2 in MS patients. We have observed a correlation between KIR2DL2 promoter methylation, leading to KIR2DL2 gene silencing, and a better MS prognosis. Altogether these data provide a novel point of view in the landscape of the immune response toward HHVs infection in MS disease, in particular it will be possible to exploit KIR2DL2 receptor expression as a target for modulating NK cell activation.File | Dimensione | Formato | |
---|---|---|---|
Tesi Silvia Bolzani Dottorato SBB ciclo 29.pdf
Open Access dal 10/03/2020
Descrizione: Tesi
Tipologia:
Tesi di dottorato
Dimensione
15.04 MB
Formato
Adobe PDF
|
15.04 MB | Adobe PDF | Visualizza/Apri |
I documenti in SFERA sono protetti da copyright e tutti i diritti sono riservati, salvo diversa indicazione.