Title

Investigating the pathological role of FGL2 in human microglia

Alternative Title

인간 미세아교세포에서 FGL2의 병리학적 역할 연구

DGIST Authors

Seunghee Lee ; Jinsoo Seo ; Seong-Woon Yu

Advisor

서진수

Co-Advisor(s)

Seong-Woon Yu

Issued Date

2024

Awarded Date

2024-02-01

Citation

Seunghee Lee. (2024). Investigating the pathological role of FGL2 in human microglia. doi: 10.22677/THESIS.200000724118

Type

Thesis

Description

Alzheimer's disease;FGL2;microglia;ApoE4;SNP (rs73375428);thrombin;autophagy

Table Of Contents

List of Contents
Abstract i
List of contents ii
List of tables and figures iv

Ⅰ. Introduction 1
1.1 Integrative analysis of eQTL and GWAS on Korean population 1
1.2 Fibrinogen-like protein 2 (FGL2) 2
1.2.1 The prothrombinase domain of FGL2 2
1.2.2 The immunosuppression domain of FGL2 3

ⅠI. Materials and Methods 5
2.1 Cell Culture 5
2.1.1 Human-induced pluripotent stem cell (hiPSC) 5
2.1.2 Immortalized Human microglia cell line (hMic) 5
2.2 Lentivirus packaging and purification 6
2.3 Antibodies 7
2.4 RNA extraction and cDNA synthesis 7
2.5 Quantitative real-time PCR 7
2.6 Aβ42 uptake 8
2.7 Immunocytochemistry 8
2.8 Immunoblotting 9
2.9 Thrombin activity assay 9
2.10 Statistical analysis 10

ⅠII. Results 11
3.1 Generation of isogenic hiPSCs using CRISPR-Cas9 system 11
3.2 Generation of isogenic hMic using CRISPR-Cas9 system 14
3.3 High expression of FGL2 protein in SNP (rs73375428) major homozygote hiPSC and ApoE4 hMic 17
3.4 Thrombin activity and Aβ42 uptake ability after drug treatment in ApoE3 and ApoE4 hMic 20
3.5 Autophagy in ApoE3 and ApoE4 hMic 25
3.6 Modulating FGL2 expression in ApoE3 and ApoE4 hMic using a viral-mediated inducible CRISPR activation/interference system 28
3.7 FGL2 overexpression and its impact on autophagy in ApoE3 hMic 33
3.8 Argatroban rescued impaired Aβ42 uptake in FGL2 Overexpressing ApoE3 hMic 37
IV. Discussion 41
V. References 44
Abstract in Korean 47

List of tables and figures
Table 1. Information of cell lines used in the research
Figure 1. Generation of isogenic SNP (rs73375428) homozygote hiPSCs
Figure 2. Generation of isogenic ApoE4 hMic
Figure 3. High expression of FGL2 in SNP (rs73375428) major homozygote hiPSC and ApoE4 hMic
Figure 4. Enhancement of Aβ42 uptake in ApoE4 hMic by Argatroban despite the similarity in thrombin activity between ApoE3 and ApoE4 hMic
Figure 5. No significant difference in autophagy between ApoE3 and ApoE4 hMic
Figure 6. Insignificant changes in FGL2 protein expression despite the mild modification of FGL2 transcripts using the CRISPRa/i system
Figure 7. FGL2 overexpression induced autophagy impairment in ApoE3 hMic
Figure 8. Restoration of FGL2 overexpression-induced impairment of Aβ42 uptake in hMic by Argatroban

URI

http://hdl.handle.net/20.500.11750/48055
http://dgist.dcollection.net/common/orgView/200000724118

DOI

10.22677/THESIS.200000724118

Degree

Master

Department

Department of Brain Sciences

Publisher

DGIST

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