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postgraduate thesis: Possible role(s) of IFN regulation on inflammasome activity via STAT1 and autophagy

TitlePossible role(s) of IFN regulation on inflammasome activity via STAT1 and autophagy
Authors
Issue Date2016
PublisherThe University of Hong Kong (Pokfulam, Hong Kong)
Citation
Pei, L. S. [貝念祖]. (2016). Possible role(s) of IFN regulation on inflammasome activity via STAT1 and autophagy. (Thesis). University of Hong Kong, Pokfulam, Hong Kong SAR.
AbstractInflammasomes are cytosolic multiprotein complex responsible for mediating inflammatory response to different pathogens, danger- and pathogen-related stimuli, thereby, providing innate immunity to our body. NLRP3 inflammasome belongs to the NLR inflammasome family and it is tightly regulated as other inflammasomes. Interferon (IFN) is one of the NLRP3 inflammasome regulators reported in the literature, however, the exact molecular mechanism remains unclear, especially in human macrophages. On the other hand, autophagy, which is a self degradative process important for energy conservation at times of nutrient deprivation and maintaining homeostasis, is another mechanism for regulating NLRP3 inflammasome activity. Our results showed that both autophagy activation and inhibitory effect of IFN on NLRP3 inflammasome operate concurrently in human monocytes derived macrophages (MDM). We hypothesize that IFN and its downstream signaling pathways are important in autophagy induction and downregulation of NLRP3 inflammasome. Since STAT1 was reported to induce autophagy machinery and the fact that STAT1 directly downstream to IFN in JAK/STAT pathway, we further hypothesize STAT1 is a potential candidate mediating the IFN inhibitory effect of IFN on NLRP3 inflammasome activity via autophagy. In this study, cytosolic LC3-II : LC3-I ratio was used as a quantitative index of autophagy activation while IL-1β secretion, caspase-1 activity and apoptosis associated speck-like protein containing a CARD (ASC) speck formation were used as measures of NLRP3 inflammasome activity. Our results demonstrated that upon both type I and type II IFN stimulation, autophagy activation was accompanied by a concurrent decrease in IL-1β secretion and caspase-1 activity in human MDM. To prove the involvement of STAT1 in autophagy activation by IFN, wild-type (WT), gain of function and loss of function STAT1 plasmid were transfected into U3C cells, a STAT1-deficient cells line. Decreased autophagy activation was shown in U3C cells transfected with loss of function STAT1 plasmid when compared to that of WT. Furthermore, transfection of STAT1 small interfering RNA (siRNA) into THP-1 cells showed a decrease in LC3-II : LC3-I ratio when compared to that of control siRNA transfected counterpart. These results demonstrated the importance of STAT1 in autophagy activation. Besides, decreased autophagy activation was also observed upon treatment with JAK1/2 inhibitor Ruxolitinib in NLRP3 inflammasome-activated MDM. All these results demonstrated the critical role of STAT1 in IFN-stimulated autophagy activation. Further effort is necessary to find out how STAT1 mediates autophagy activation. The elucidation of the detailed molecular pathways on the inhibitory effect of IFN and JAK-STAT pathway may provide opportunities to develop therapeutic strategies toward autoinflammatory diseases.
DegreeDoctor of Philosophy
SubjectMolecular aspects - Inflammation
Interferon
Dept/ProgramPaediatrics and Adolescent Medicine
Persistent Identifierhttp://hdl.handle.net/10722/270248

 

DC FieldValueLanguage
dc.contributor.authorPei, Lim-cho, Steven-
dc.contributor.author貝念祖-
dc.date.accessioned2019-05-23T02:26:19Z-
dc.date.available2019-05-23T02:26:19Z-
dc.date.issued2016-
dc.identifier.citationPei, L. S. [貝念祖]. (2016). Possible role(s) of IFN regulation on inflammasome activity via STAT1 and autophagy. (Thesis). University of Hong Kong, Pokfulam, Hong Kong SAR.-
dc.identifier.urihttp://hdl.handle.net/10722/270248-
dc.description.abstractInflammasomes are cytosolic multiprotein complex responsible for mediating inflammatory response to different pathogens, danger- and pathogen-related stimuli, thereby, providing innate immunity to our body. NLRP3 inflammasome belongs to the NLR inflammasome family and it is tightly regulated as other inflammasomes. Interferon (IFN) is one of the NLRP3 inflammasome regulators reported in the literature, however, the exact molecular mechanism remains unclear, especially in human macrophages. On the other hand, autophagy, which is a self degradative process important for energy conservation at times of nutrient deprivation and maintaining homeostasis, is another mechanism for regulating NLRP3 inflammasome activity. Our results showed that both autophagy activation and inhibitory effect of IFN on NLRP3 inflammasome operate concurrently in human monocytes derived macrophages (MDM). We hypothesize that IFN and its downstream signaling pathways are important in autophagy induction and downregulation of NLRP3 inflammasome. Since STAT1 was reported to induce autophagy machinery and the fact that STAT1 directly downstream to IFN in JAK/STAT pathway, we further hypothesize STAT1 is a potential candidate mediating the IFN inhibitory effect of IFN on NLRP3 inflammasome activity via autophagy. In this study, cytosolic LC3-II : LC3-I ratio was used as a quantitative index of autophagy activation while IL-1β secretion, caspase-1 activity and apoptosis associated speck-like protein containing a CARD (ASC) speck formation were used as measures of NLRP3 inflammasome activity. Our results demonstrated that upon both type I and type II IFN stimulation, autophagy activation was accompanied by a concurrent decrease in IL-1β secretion and caspase-1 activity in human MDM. To prove the involvement of STAT1 in autophagy activation by IFN, wild-type (WT), gain of function and loss of function STAT1 plasmid were transfected into U3C cells, a STAT1-deficient cells line. Decreased autophagy activation was shown in U3C cells transfected with loss of function STAT1 plasmid when compared to that of WT. Furthermore, transfection of STAT1 small interfering RNA (siRNA) into THP-1 cells showed a decrease in LC3-II : LC3-I ratio when compared to that of control siRNA transfected counterpart. These results demonstrated the importance of STAT1 in autophagy activation. Besides, decreased autophagy activation was also observed upon treatment with JAK1/2 inhibitor Ruxolitinib in NLRP3 inflammasome-activated MDM. All these results demonstrated the critical role of STAT1 in IFN-stimulated autophagy activation. Further effort is necessary to find out how STAT1 mediates autophagy activation. The elucidation of the detailed molecular pathways on the inhibitory effect of IFN and JAK-STAT pathway may provide opportunities to develop therapeutic strategies toward autoinflammatory diseases. -
dc.languageeng-
dc.publisherThe University of Hong Kong (Pokfulam, Hong Kong)-
dc.relation.ispartofHKU Theses Online (HKUTO)-
dc.rightsThe author retains all proprietary rights, (such as patent rights) and the right to use in future works.-
dc.rightsThis work is licensed under a Creative Commons Attribution-NonCommercial-NoDerivatives 4.0 International License.-
dc.subject.lcshMolecular aspects - Inflammation-
dc.subject.lcshInterferon-
dc.titlePossible role(s) of IFN regulation on inflammasome activity via STAT1 and autophagy-
dc.typePG_Thesis-
dc.description.thesisnameDoctor of Philosophy-
dc.description.thesislevelDoctoral-
dc.description.thesisdisciplinePaediatrics and Adolescent Medicine-
dc.description.naturepublished_or_final_version-
dc.date.hkucongregation2016-
dc.identifier.mmsid991044104201803414-

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