• BMB Reports
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• v.57 no.1
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• pp.60-65
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• 2024

The CRISPR-Cas9 system has significantly advanced regenerative medicine research by enabling genome editing in stem cells. Due to their desirable properties, mesenchymal stem cells (MSCs) have recently emerged as highly promising therapeutic agents, which properties include differentiation ability and cytokine production. While CRISPR-Cas9 technology is applied to develop MSC-based therapeutics, MSCs exhibit inefficient genome editing, and susceptibility to plasmid DNA. In this study, we compared and optimized plasmid DNA and RNP approaches for efficient genome engineering in MSCs. The RNP-mediated approach enabled genome editing with high indel frequency and low cytotoxicity in MSCs. By utilizing Cas9 RNPs, we successfully generated B2M-knockout MSCs, which reduced T-cell differentiation, and improved MSC survival. Furthermore, this approach enhanced the immunomodulatory effect of IFN-r priming. These findings indicate that the RNP-mediated engineering of MSC genomes can achieve high efficiency, and engineered MSCs offer potential as a promising therapeutic strategy.

• Korean Journal of Radiology
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• v.23 no.6
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• pp.638-648
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• 2022

Objective: The study aimed to investigate the diurnal variation in cervical disc hydration and its relationship with cervical degeneration. Materials and Methods: C3-C7 discs of 86 prospectively enrolled participants (37 males, 49 females; mean age ± standard deviation, 23.5 ± 2.5 years) were assessed using T2 mapping in the morning and evening. All discs were stratified by Miyazaki grade or C2-C7 Cobb angle and T2 values (T2). The degree of diurnal T2 variation (T2-DDV), defined as (morning T2 - evening T2)/morning T2 x 100%, was measured for the entire disc, annulus fibrosus (AF), nucleus pulposus (NP), and endplate zones. Results: T2 of the entire disc decreased significantly after the daytime load (p < 0.001), with a T2-DDV of 13.3% for all discs and 16.0%, 12.2%, and 13.0% for healthy (grade I), mild degenerative (grade II), and advanced degenerative (grade III/IV) discs, respectively. T2 of regional NPs and AFs decreased significantly from morning to evening (p ≤ 0.049) except in the healthy anterior inner AF (p = 0.092). Compared with healthy discs, mild degenerative discs displayed lower T2 and T2-DDV in regional NPs (p < 0.001). Advanced degenerative discs showed higher T2-DDV in the anterior inner AF compared with healthy discs (p = 0.050). Significant diurnal T2 changes in the endplate zones were observed only in healthy discs (p = 0.013). Cervical discs in the low Cobb angle group showed higher T2-DDV in the anterior AFs and anterior NP and lower T2-DDV in the posterior AF than those in the high Cobb angle group (p ≤ 0.041). Conclusion: This study characterized the diurnal variation in hydration of the cervical discs as assessed using T2 mapping and revealed early chemo-mechanical coupling dysfunction in degenerating discs. Cervical sagittal alignment on MRI can affect the diurnal stress patterns of the cervical discs. T2 mapping is sensitive to disc biomechanical dysfunction and offers translational potential from biomechanical research to clinical application.

• Journal of Dental Rehabilitation and Applied Science
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• v.34 no.4
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• pp.280-289
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• 2018

Purpose: The purpose of this randomized clinical trial is to determine whether implant length and the crown-to-implant (C/I) ratio influence implant stability and peri-implant marginal bone loss (MBL). Materials and Methods: 46 patients with single tooth missing in the posterior molar region of the mandible were included in this study. 19 implants (CMI IS-III $active^{(R)}$ long implant) of 5.0 mm diameter and 10 mm length were installed for the control group, while 27 implants (CMI IS-III $active^{(R)}$ short implant) of 5.5 mm diameter and 6.6, 7.3 or 8.5 mm length were placed for the experimental group. Each implant was inserted and immediately loaded using the digitally pre-fabricated surgical template and provisional restoration. The CAD-CAM monolithic zirconia crown was fabricated at 3 months after the surgery as a definitive restoration. The ISQ value and the MBL was measured at 48 weeks after the surgery. The correlation between the C/I ratio, MBL, and secondary implant stability was analyzed. Results: Successful results in terms of ISQ and MBL were achieved with both groups. There was no significant difference between the groups in terms of ISQ values and MBL at 48 weeks after the surgery (P > 0.05). No significant correlation was found between the C/I ratio and secondary stability as well as the C/I ratio and the MBL (P > 0.05). Conclusion: The influence of C/I ratio in both groups was not shown on the stability nor the marginal bone loss in implants supporting single crown of the mandible. Short implant could be a preferable alternative option in the reduced bone height mandible under the limited condition despite its higher C/I ratio.

• Journal of Dental Rehabilitation and Applied Science
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• v.34 no.4
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• pp.270-279
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• 2018

Purpose: The aim of this study was to assess the patients' perception, acceptance, and preference of the difference between a conventional impression and digital impression through questionnaire survey. Materials and Methods: Thirteen (6 male, 7 female) subjects who experienced both digital and conventional impression at the same day were enrolled in this study. Conventional impression were taken with polyvinylsiloxane and digital impression were performed using a newly developed intra-oral scanner. Immediately after the two impressions were made, a survey was conducted with the standardized questionnaires consisting of the following three categories; 1) general dental treatment 2) satisfaction of conventional impression 3) satisfaction of digital impression. The perceived source of satisfaction was evaluated using Likert scale. The distribution of the answers was assessed by percentages and statistical analyses were performed with the paired t-test, and P < 0.05 was considered significant. Results: There were significant differences of the overall satisfaction between two impression methods (P < 0.05). Digital impression showed high satisfaction in less shortness of breath and odor to participants compared to conventional impression. The use of an oral scanner resulted in a discomfort of TMJ due to prolonged mouth opening and in lower score of the scanner tip size. Conclusion: It was confirmed that the preference for the digital impression using intraoral scanner is higher than the conventional impression. Most survey participants said they would recommend the digital impression to others and said they preferred it for future prosthetic treatment.

• The Journal of Korean Academy of Prosthodontics
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• v.59 no.2
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• pp.153-163
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• 2021

Purpose: The aim of this study was to evaluate the effects of four surface treatment methods to improve zirconia roughness and three types of resin cement on the shear bond strength (SBS). Materials and methods: A total of 120 zirconia blocks were randomly divided into four surface treatments: non-treatment (Control), airborne-particle abrasion (APA) with 50 ㎛ Al2O3 (APA50), APA with 125 ㎛ Al2O3 (APA125), and ZrO2 slurry (ZA). Three resin cements (Panavia F 2.0, Superbond C&B, and Variolink N) were applied to the surface-treated zirconia specimens. All specimens were subjected to SBS testing using a universal testing machine. The surface of the representative specimens of each group was observed by scanning electron microscope (SEM). SBS data were analyzed with oneway ANOVA, two-way ANOVA test and post-hoc Tukey HSD Test (α=.05). Results: In the surface treatment method, APA125, APA50, ZA, and Control showed high shear bond strength in order, but there was no significant difference between APA125 and APA50 (P>.05). Also, ZA showed significantly higher shear bond strength than Control (P<.05). In the resin cement type, Panavia F 2.0, Superbond C&B, and Variolink N showed significantly higher shear bond strength in order (P<.05). In SEM images, the zirconia surfaces of the APA50 and APA125 showed quite rough and irregular shapes, and the zirconia surface of the ZA was observed small irregular porosity and rough surfaces. Conclusion: APA and ZrO2 slurry were enhanced the surface roughness of zirconia, and Panavia F 2.0 containing MDP showed the highest shear bond strength with zirconia.

• Journal of Life Science
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• v.34 no.2
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• pp.86-93
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• 2024

Since replication of plasmids must be strictly controlled, plasmids that generally perform rolling circle replication generally maintain a constant copy number by strictly controlling the replication initiator Rep at the transcriptional and translational levels. Plasmid pJB01 contains three orfs (copA, repB, repC or repABC) consisting of a single operon. From analysis of amino acid sequence, pJB01 CopA was homologous to the Cops, as a copy number control protein, of other plasmids. When compared with a CopG of pMV158, CopA seems to form the RHH (ribbon-helix-helix) known as a motif of generalized repressor of plasmids. The result of gel mobility shift assay (EMSA) revealed that the purified fusion CopA protein binds to the operator region of the repABC operon. To examine the functional role of CopA on transcriptional level, 3 point mutants were constructed in coding frame of copA such as CopA R16M, K26R and E50V. The repABC mRNA levels of CopA R16M, K26R and E50V mutants increased 1.84, 1.78 and 2.86 folds more than that of CopA wt, respectively. Furthermore, copy numbers owing to mutations in three copA genes also increased 1.86, 1.68 and 2.89 folds more than that of copA wt, respectively. These results suggest that CopA is the transcriptional repressor, and lowers the copy number of pJB01 by reducing repABC mRNA and then RepB, as a replication initiator.

• Journal of radiological science and technology
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• v.37 no.1
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• pp.37-47
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• 2014

The purpose of this study was to analyze translational and rotational adjustments during automatic rigid image-registration by using different control parameters for a total of five groups on TomoTherapy (Accuray Inc, Sunnyvale, CA, USA). We selected a total of 50 patients and classified them in five groups (brain, head-and-neck, lung, abdomen and pelvic) and used a total of 500 megavoltage computed tomography (MVCT) image sets for the analysis. From this we calculated the overall mean value(M) for systematic and random errors after applying the different control parameters. After randomization of the patients into the five groups, we found that the overall mean value varied according to three techniques and resolutions. The deviation for the lung, abdomen and pelvic groups was approximately greater than the deviation for the brain and head-and-neck groups in all adjustments. Overall, using a "full-image" produces smaller deviations in the rotational adjustments. We found that rotational adjustment has deviations with distinctly different control parameters. We concluded that using a combination of the "full-image" technique and "standard" resolution will be helpful in assisting with patients' repositioning and in correcting for set-up errors prior to radiotherapy on TomoTherapy.

• Korean Journal of Microbiology
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• v.46 no.1
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• pp.21-26
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• 2010

As the causative agent of Anthrax, Bacillus anthracis causes an acute fatal disease in herbivores such as cattle, sheep, and horses as well as humans. The therapeutics and prevention of anthrax currently available are based on antibiotics and the live attenuated vaccine strains, which may be problematic due to the emergency of antibiotic resistant strains or residual virulence in those vaccine strains. Therefore, it has been required to develop novel therapeutics and vaccines which are safer and applicable to humans. Recently, the development of the multivalent vaccine targeting both spores and vegetative cells of B. anthracis along with anthrax toxin has been reported. In our attempts to screen potential candidates for those multivalent vaccines, the whole genomic expression library of B. anthracis was constructed in this study. To the end, the partial digests of the genomic DNA from B. anthracis (ATCC 14578) with Sau3AI were ligated with the inducible pET30abc expression vectors, resulting in approximately $1{\times}10^5$ clones in E. coli BL21(DE3). The redundancy test by DNA nucleotide sequencing was performed for the randomly selected 111 clones and found 56 (50.5%) B. anthracis genes, 17 (15.3%) vector sequences, and 38 (34.2%) unknown genes with no sequence homology by BLAST. An inducible expression of the recombinant proteins was confirmed by Western blot. Interestingly, some clones could react with the antiserum against B. anthracis. These results imply that the whole genomic library constructed in this study can be applied for analyzing the immunomes of B. anthracis.

• Journal of Life Science
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• v.23 no.11
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• pp.1397-1403
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• 2013

Fructus Sophorae, the dried ripe fruit of Styphnolobium japonicum (L.), is an herbal ingredient used in traditional Oriental medicine. This study was carried out to investigate the effects of Fructus Sophorae extracts (FSE) on immune modulation in a murine RAW 264.7 macrophage model. As immune response parameters, the production of prostaglandin $E_2$ ($PGE_2$) and tumor necrotic $factor-{\alpha}$ ($TNF-{\alpha}$) were evaluated. Our data revealed that FSE increased the macrophage activation and the production of $PGE_2$ and $TNF-{\alpha}$, which was consistently correlated with upregulation of cyclooxygenase-2 (COX-2) and $TNF-{\alpha}$ expression at both transcriptional and translational levels. On comparative cytokine protein array, FSE significantly increased several cytokines, which was associated with phosphorylation of mitogen- activated protein kinases (MAPKs), including extracellular signal-regulated kinase (ERK), p38 MAPK and c-Jun N-terminal kinase (JNK), and Akt in RAW 264.7 cells. However, each inhibitor of these molecules attenuated the FSE-induced $PGE_2$ production. These results indicate that FSE activated macrophages through the activation of MAPKs and phosphatidylinositol-3-kinase (PI3K)/Akt signaling pathways in RAW 264.7 macrophages. These findings suggest that FSE may provide a promising source of an immunoenhancing agent.

• Clinical and Experimental Pediatrics
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• v.49 no.4
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• pp.431-438
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• 2006

Purpose : Insulin-like growth factor binding protein(IGFBP)-3 has been known as a tumor suppressor gene, and its anti-tumor function was divided into insulin-like growth factor(IGF)-dependent and IGF-independent mechanism. In IGF-independent mechanism, IGFBP-3 directly interacts with a cell without binding of IGFs, becoming an interesting object in oncology. Several studies demonstrate that one of the well-known tumor suppressor genes, p53, induces directly IGFBP-3 transcription, and the increment of IGFBP-3 expression induces apoptosis of many cancer cells. Recently, the anti-tumor mechanisms of IGFBP-3 have been reported, but post-translational modification of IGFBP-3 and its anti-tumor mechanism are not well known. In this study, we examined whether p53 regulated the glycosylation of IGFBP-3, and analysed the meaning of IGFBP-3 glycosylation related to the apoptosis of cancer cell. Methods : The p53-mutated status of MDA-MB-231 human breast cancer cells was used in this experiment. The expression and glycosylation of IGFBP-3 were tested by Western blot analysis after infection of adenovirus mediated Ad/p53 and/or Ad/IGFBP-3. Results : Ad/p53 infected cells resulted in growth retardation and the induced apoptosis. p53 induced direct expression and glycosylation of IGFBP-3. The increase of glcosylated IGFBP-3 was able to promote cellular apoptosis, and the glycosylation of IGFBP-3 was more activated by the double treatment of Ad/p53 and Ad/IGFBP-3. Conclusion : From this study, the anti-tumor activity of IGFBP-3 was shown to improve the stabilization of IGFBP-3 through the increment of glycosylation of IGFBP-3 by p53. This result suggests that the combined gene therapy of p53 and IGFBP-3 may appropriate treatment of cancer.