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A Novel DNA-PK/PLK1/STING/IRF3 Axis Regulates Normal Mitotic Progression
No full textDoctor1. INTRODUCTION 1
2. MATERIALS AND METHODS 10
2.1. Cell culture 10
2.2. Antibodies 10
2.3. Synchronization and drug treatment 11
2.4. Plasmids and transfection experiments 12
2.5. Knock-down experiments 12
2.6. Immunoblotting 14
2.7. cGAMP ELISA 15
2.8. Native-PAGE 15
2.9. Time-lapse analysis 16
2.10. 2D Blue Native-PAGE 16
2.11. Immunocytochemistry 17
2.12. Immunoprecipitation 17
2.13. In-vitro kinase assay 18
2.14. Quantitative RT-PCR 19
2.15. Mitotic index 19
2.16. Statistical analysis 20
3. RESULTS 21
3.1. IRF3 is phosphorylated during mitosis in a cGAS-independent but STING-dependent manner 21
3.2. Phosphorylation of IRF3 is required for normal mitotic progression 28
3.3. STING and IRF3 dimerization is a pre-requisite during mitosis 35
3.4. DNA-PKcs activity is required for STING and IRF3 phosphorylation during mitosis 39
3.5. PLK1 is the major mitotic kinase responsible for IRF3 phosphorylation during mitosis 47
3.6. STING, IRF3, DNA-PK and PLK1 form a complex during mitosis where STING acts as a mediator 53
3.7. Recruitment of PLK1 and IRF3 to the DNA-PK/STING complex requires DNA-PK activity 62
3.8. Ku70 and Ku80 may play different roles in the activation of STING/IRF3 axis in a cell-dependent manner 66
4. DISCUSSION 71
5. REFERENCES 7
YAP이 Ku70 또는 MRE11에 의해 매개되는 초기 DNA 손상 인식 과정에 관여한다
No full textMasterⅠ. Introduction 1
ⅠⅠ. Materials and Methods 11
1. Cell culture 11
2. Plasmid DNA transfection 11
3. CRISPR/Cas9 system for generating knock out cell lines 11
4. TA cloning 12
5. Western blot 12
6. Co-immunoprecipitation (Co-IP) 13
7. GST-fusion protein purification 13
8. In vitro GST pull-down assay 14
9. Subcellular fractionation 14
10. Molecular docking model 15
11. Live-cell imaging with laser micro-irradiation 15
12. Immunocytochemistry 16
13. Lentivirus production and generation of stable knockdown cell lines 16
14. Gene knockdown using siRNA 17
15. DNA repair reporter assay 17
16. Clonogenic assay 17
17. Antibodies 18
18. Chemicals 18
ⅠⅠⅠ. Results 19
1. YAP directly interacts with Ku70 19
2. The WW2 domain of YAP is the key mediator of its interaction with Ku70 22
3. The interaction between YAP and Ku70 is enhanced upon DNA damage 25
4. YAP depletion attenuates DDR and DSB repair efficiency 28
5. YAP depletion induces genomic instability 36
ⅠⅤ. Discussion 39
Ⅴ. References 43
국문 초록 5
Genome-Wide Association Study to Identify Genetic Factors Linked to HBV Reactivation Following Liver Transplantation in HBV-Infected Patients
No full textThis study utilized a genome-wide association study (GWAS) to investigate the genetic variations linked to the risk of hepatitis B virus (HBV) reactivation in patients who have undergone liver transplantation (LT), aiming to enhance understanding and improve clinical outcomes. Genotyping performed on a selected patients from the Korean Organ Transplantation Registry (KOTRY) data using high-throughput platforms with the Axiom Korea Biobank array 1.1. The discovery cohort included 21 patients who experienced HBV reactivation (cases) and 888 patients without HBV reactivation (controls) following LT. The replication cohort consisted of 5 patients with HBV reactivation (cases) and 312 patients without HBV reactivation (controls) after LT. Additive logistic regression analysis was conducted using PLINK software ver 1.9, with adjustments for age and gender. The GWAS findings from the discovery cohort were validated using the replication cohort. The GWAS identified several single-nucleotide polymorphisms (SNPs) in the RGL1, CDCA7L, and AQP9 genes that were significantly linked to HBV reactivation after LT, with genome-wide significance thresholds set at p < 10−7. Down-regulation of RGL1 cDNAs was observed in primary duck hepatocytes infected with duck HBV. Overexpression of CDCA7L was found to promote hepatocellular carcinoma cell proliferation and colony formation, whereas knocking down CDCA7L inhibited these processes. Additionally, the absence of AQP9 triggered immune and inflammatory responses, leading to mild and scattered liver cell pyroptosis, accompanied by compensatory liver cell proliferation. This study provides critical insights into the genetic factors influencing HBV reactivation after LT, identifying significant associations with SNPs in RGL1, CDCA7L, and AQP9. These findings hold promise for developing predictive biomarkers and personalized management strategies to improve outcomes for HBV-infected LT recipients
SB16 versus reference denosumab in postmenopausal women with osteoporosis: 18-month outcomes of a phase III randomized clinical trial
No full textPurpose: This study evaluated the efficacy, safety, pharmacodynamics (PD), pharmacokinetics (PK), and immunogenicity of SB16 versus reference denosumab (DEN) up to 18 months in postmenopausal osteoporosis (PMO) patients, and assessed outcomes after switching from DEN to SB16 compared to those who continued with DEN or SB16. Methods: 457 PMO patients were initially randomized, with 407 re-randomized at Month 12 to either continue DEN (DEN+DEN), switch to SB16 (DEN+SB16), or continue SB16 (SB16 + SB16) through Month 18. Efficacy was assessed by the percent change from baseline in bone mineral density (BMD) at the lumbar spine, total hip, and femoral neck. Safety, PD, PK, and immunogenicity were evaluated throughout the study period. Results: Mean percent changes from baseline in lumbar spine, total hip, and femoral neck BMD at Month 18 were comparable across treatment groups, indicating comparable efficacy between SB16 and DEN. The mean percent change in lumbar spine BMD was 6.8 % (SB16 + SB16), 6.2 % (DEN+SB16), and 6.8 % (DEN+DEN). Total hip BMD increased by 4.4 %, 3.5 %, and 4.0 %, and femoral neck BMD by 3.4 %, 3.1 %, and 2.7 % for SB16 + SB16, DEN+SB16, and DEN+DEN, respectively. Safety profiles were similar among groups, with no new safety concerns identified after switching. Only one patient in the DEN+SB16 group developed non-neutralizing anti-drug antibodies by Month 18, indicating a low immunogenicity risk for SB16. Conclusion: Switching from DEN to SB16 demonstrated comparable efficacy, safety, PD, PK, and immunogenicity in PMO patients relative to those who continued DEN. SB16 was well tolerated over 18 months, demonstrating comparable outcomes to DEN
Automated quantitative pupillometry as a predictor for transtentorial brain herniation in patients with malignant acute ischemic stroke
No full textBrain herniation can be a life-threatening condition, resulting in poor prognosis and higher fatality rates. We examined whether quantitative characteristics of sequential pupillary light reflex (PLR) could serve as biomarkers for identifying brain herniation in fatal acute stroke cases with anterior circulation involvement admitted to neurological intensive care unit (Neuro-ICU). Automatic pupillometer assessed PLR automatically every 4–6 hours, measuring eight specific features: NPi (Neurological pupil index) score, initial resting and constriction pupil size, constriction change, constriction velocity, constriction latency, and dilation velocity. Generalized estimating equations were used to analyze the main effects of assessment time (3-to-0 hours, just before brain herniation, and 27-to-21 hours, considerably before) and clinical groups. The study involved 59 patients (mean age 68.8 ± 1.6 years, 23 females) divided into herniation (n = 10) and non-herniation (n = 49) groups. The herniation group exhibited significantly lower ipsilateral NPi scores at 3-to-0 hours (1.80 ± 0.44, p < 0.0001) compared to 27-to-21 hours (4.26 ± 2.21). Additionally, the herniation group had a larger ipsilateral pupil size at constriction at 3-to-0 hours (4.01 ± 0.40 mm) compared to 27-to-21 hours (2.11 ± 0.17 mm). Specifically, at 3-to-0 hours, the herniation group had lower NPi scores (1.80 ± 0.44 vs. 3.97 ± 0.13, p < 0.0001) and larger pupil size at constriction (4.01 ± 0.04 mm vs. 2.90 ± 0.10 mm, p = 0.007) compared to the non-herniation group. These findings suggest that evaluating PLR characteristics can aid in the early identification of brain herniation, facilitating timely triage and appropriate surgical management
Rifaximin treatment in patients with severe alcoholic hepatitis: A multicenter, randomized controlled, open-label, pilot trial
No full textINTRODUCTION AND OBJECTIVES: The short-term mortality of severe alcoholic-associated hepatitis (SAH) is high, but there are no effective treatments to improve short-term mortality other than corticosteroids. This study investigated the effects of adding rifaximin to standard treatment in patients with SAH. MATERIAL AND METHODS: In this randomized controlled open-label trial, patients with SAH (Maddrey's discriminant function>/=32) were randomized to the rifaximin or control group. Patients were simultaneously treated with corticosteroid or pentoxifylline as standard treatment for 4 weeks. Randomization was stratified by SAH treatment. RESULTS: A total of 50 patients were enrolled in this study (29 in the control group and 21 in the rifaximin group). The mean Model for End-stage Liver Disease (MELD) scores were 24.4 and 27.5 in the control and rifaximin groups, respectively (P = 0.106). There were no significant differences in 6-month Liver Transplantation (LT)-free survival rate between the two groups (P = 0.502). When stratified by SAH treatment, there was no significant difference in 6-month LT-free survival rate between the control and rifaximin treatment groups (P = 0.186 in the corticosteroid group and P = 0.548 in the pentoxifylline group). There were no significant differences in the occurrence of liver-related complications between the two groups (all Ps>0.05). The MELD score was the only independent factor for 6-month LT-free survival (hazard ratio 1.188, 95 % confidence interval 1.094-1.289, P<0.001), and rifaximin was not. CONCLUSIONS: In patients with SAH, adding rifaximin to corticosteroid or pentoxifylline had no survival benefit and no preventive effect on the development of liver-related complications. The MELD score was the only significant factor for short-term mortality. CLINICAL TRIAL REGISTRATION: The study was registered on ClinicalTrials.gov (number: NCT02485106)
Comparison of the efficacy and safety of bupropion versus aripiprazole augmentation in adults with treatment-resistant depression: A nationwide cohort study in South Korea
No full textBACKGROUND: Treatment-resistant depression (TRD) affects 10-30% of patients with major depressive disorder, leading to increased comorbidities, higher mortality, and significant economic and social burdens. This study aimed to compare the efficacy and safety of bupropion and aripiprazole as augmentation therapies for TRD. METHODS: This population-based, retrospective cohort study included adults aged >/=18 years with a diagnosis of depressive disorder who met the criteria for TRD. Data were collected from a nationwide claims database in South Korea. Patients prescribed bupropion were matched 1:1 with those prescribed aripiprazole. Subgroup analyses were performed according to age. An as-treated analysis was performed as the primary analysis, and an intention-to-treat analysis was performed to identify different risk windows. The primary outcome was depression-related hospitalization, and the secondary outcomes were first-time diagnoses of movement disorder and seizure. RESULTS: A total of 5,619 patients (bupropion: n = 1,568; aripiprazole: n = 4,051) were included in this study. Bupropion was associated with lower risks of hospitalization (hazard ratio [HR]: 0.51; 95% confidence interval [CI] 0.29-0.86) and movement disorders (HR: 0.56; 95% CI 0.36-0.85) than aripiprazole. No significant difference in seizure risk (HR: 0.65; 95% CI 0.30-1.31) was observed between the two treatments. The subgroup analysis of participants aged >/=60 years revealed no significant differences in the three outcomes between the two medications. CONCLUSIONS: Bupropion augmentation is associated with a significantly lower risk of depression-related re-hospitalization and movement disorders in patients with TRD. Therefore, bupropion augmentation can be a comprehensive treatment strategy for TRD
Clinical practice guidelines for ovarian cancer: an update to the Korean Society of Gynecologic Oncology guidelines
No full textWe updated the Korean Society of Gynecologic Oncology (KSGO) practice guideline for the management of ovarian cancer as version 5.1. The ovarian cancer guideline team of the KSGO published announced the fifth version (version 5.0) of its clinical practice guidelines for the management of ovarian cancer in December 2023. In version 5.0, the selection of the key questions and the systematic reviews were based on the data available up to December 2022. Therefore, we updated the guidelines version 5.0 with newly accumulated clinical data and added 5 new key questions reflecting the latest insights in the field of ovarian cancer between 2023 and 2024. For each question, recommendation was provided together with corresponding level of evidence and grade of recommendation, all established through expert consensus
Drosophila ubiquitin-specific peptidase 14 stabilizes the PERIOD protein by regulating a ubiquitin ligase SLIMB
No full textThe circadian clock orchestrates behavior and physiology through the oscillation of key clock proteins like PERIOD (PER). Here, we investigate the role of ubiquitin-specific peptidase 14 (USP14) in modulating PER stability and circadian rhythms in Drosophila. We find that overexpression of USP14 in clock cells reduces PER protein levels without altering its mRNA levels whereas USP14 knockdown increases PER protein levels, suggesting that USP14 regulates PER post-translationally. Interestingly, despite these alterations in PER levels, neither USP14 overexpression nor knockdown significantly impacts circadian behavioral rhythms, likely because of slight effects on PER levels in small ventral lateral neurons (sLN(v)s). Further analysis shows that USP14 physically interacts with Supernumerary Limbs (SLIMB), a protein involved in PER degradation. Moreover, reducing slimb expression mitigates the effects of USP14 on PER protein stability. Mass spectrometry identifies two ubiquitination sites on PER (Lys1117 and Lys1118) critical for its degradation. Expression of PER(1117A, 1118A) mutant in per(01) background impairs circadian rhythm strength. In conclusion, this study demonstrates that Drosophila USP14 indirectly modulates PER protein stability by affecting SLIMB and highlights the critical role of specific ubiquitination sites on PER in maintaining circadian rhythms
Inability to Remove Locking Screws From the Femoral Neck System Due to Stripping of the Screwdriver Within the Locking Screw Head
No full textOBJECTIVES: The purposes of this study were to investigate the frequency of screwdriver stripping in the head of the locking screw that attaches to the side plate to the femur shaft among the patients who underwent implant removal after femoral neck system (FNS) for femoral neck fracture, to determine the risk factors for locking head screw stripping in FNS treatment of femoral neck fracture, and to suggest a surgical tip that removes FNS, which is difficult to remove due to screw stripping. DESIGN: Retrospective cohort study. SETTING: Eight Urban tertiary referral academic hospitals. PATIENT SELECTION CRITERIA: Included were patients with Orthopedic Trauma Association/Arbeitsgemeinschaft fur Osteosynthesefragen 31-B1, 31-B2, and 31-B3 femoral neck fractures who underwent surgical fixation with FNS from November 2019 to February 2023. OUTCOME MEASURES AND COMPARISONS: The frequency of locking head screw stripping of FNS during the implant removal was evaluated. RESULTS: Among the 47 patients (18 (38%) men and 29 (62%) women) who met the inclusion criteria with an average age of 59.2 years (range, 28-94 years), 13 (27.7%) experienced screwdriver stripping in the head of the distal locking screw during FNS removal surgery. A higher body mass index showed a borderline significant association with the stripping in the adjusted model (odds ratio = 1.233; 95% confidence interval: 0.988-1.539; P = 0.064). No other variables showed significant association with the stripped locking head screw ( P > 0.05). CONCLUSIONS: Stripping of the screwdriver within the head of the distal locking screw occurred in over one-quarter of cases. While a higher body mass index demonstrated a borderline significant association, none of the other variables examined showed a statistically significant relationship with the stripped locking head screw. LEVELS OF EVIDENCE: Prognostic Level III. See Instructions for Authors for a complete description of levels of evidence