Background: Iris Koreana NAKAI (IKN) is a flowering perennial plant that belongs to the Iridaceae family. In this study, we aimed to demonstrate the effects of IKN on osteoclast differentiation in vitro and in vivo. We also sought to verify the molecular mechanisms underlying its anti-osteoclastogenic effects. Methods: Osteoclasts were formed by culturing mouse bone marrow macrophage (BMM) cells with macrophage colony-stimulating factor and receptor activator of nuclear factor-κB ligand (RANKL). Bone resorption assays were performed on dentin slices. mRNA expression levels were analyzed by quantitative polymerase chain reaction. Western blotting was performed to detect protein expression or activation. Lipopolysaccharide (LPS)-induced osteoclast formation was performed using a mouse calvarial model. Results: In BMM cultures, an ethanol extract of the root part of IKN suppressed RANKL-induced osteoclast formation and bone resorptive activity. In contrast, an ethanol extract of the aerial parts of IKN had a minor effect on RANKL-induced osteoclast formation. Mechanistically, the root part of IKN suppressed RANKL-induced p38 mitogen-activated protein kinase (MAPK) activation, effectively abrogating the induction of c-Fos and nuclear factor of activated T cells 1 (NFATc1) expression. IKN administration decreased LPS-induced osteoclast formation in a calvarial osteolysis model in vivo. Conclusions Our study suggested that the ethanol extract of the root part of IKN suppressed osteoclast differentiation and function partly by downregulating the p38 MAPK/c-Fos/NFATc1 signaling pathways. Thus, the root part

BACKGROUND: Osteoclasts are the only cell type capable of breaking down bone matrix, and its excessive activation is responsible for the development of bone-destructive diseases. Euphorbia lathyris L. (ELL) is an herbal plant that belongs to the Euphorbiaceae family. This study investigated the effects of the methanol extract of the aerial part of ELL on receptor activator of nuclear factor-kappa B ligand (RANKL)-induced osteoclast formation and signaling pathways. METHODS: Osteoclasts were formed by co-culturing mouse bone marrow with osteoblasts or by culturing mouse bone marrow-derived macrophages (BMMs) with macrophage colony-stimulating factor (M-CSF) and RANKL. Bone resorption assays were performed using dentine slices. The expression level of mRNA was analyzed by real-time polymerase chain reaction (PCR) or reverse transcription (RT)-PCR. Western blotting assays were performed to detect the expression or activation level of proteins. RESULTS: ELL inhibited RANKL-induced osteoclast formation without cytotoxicity. Furthermore, the RANKL-stimulated bone resorption was diminished by ELL. Mechanistically, ELL blocked the RANKL-triggered p38 mitogen-activated protein kinase (MAPK) phosphorylation, which resulted in the suppression of the expression of c-Fos and nuclear factor of activated T cells (NFATc1). In osteoblasts, ELL had little effect on the mRNA expression of RANKL and osteoprotegerin (OPG). CONCLUSIONS: The present data suggest that ELL has an inhibitory effect on osteoclast differentiation and function via downregulation of the p38/c-Fos/NFATc1 signaling pathways. Thus, ELL could be useful for the treatment of bone diseases associated with excessive bone resorption.
Animals ; Blotting, Western ; Bone Diseases ; Bone Marrow ; Bone Matrix ; Bone Resorption ; Dentin ; Down-Regulation ; Euphorbia* ; Euphorbiaceae ; Humans ; Macrophage Colony-Stimulating Factor ; Macrophages ; Methanol ; Mice ; Osteoblasts ; Osteoclasts* ; Osteoprotegerin ; p38 Mitogen-Activated Protein Kinases ; Phosphorylation ; Plants ; Protein Kinases ; RANK Ligand ; Real-Time Polymerase Chain Reaction ; Reverse Transcription ; RNA, Messenger ; T-Lymphocytes

OBJECTIVES: Osteoclasts (OCs) are bone-resorbing multinucleated cells derived from hematopoietic progenitors of the monocyte-macrophage lineage. OC precursors, such as bone marrow-derived macrophages (BMMs), are formed in the presence of macrophage colony-stimulating factor (M-CSF) and differentiate into OCs in response to M-CSF and receptor activator of nuclear factor kappaB ligand (RANKL). In this study, we investigated the role of mixed lineage kinases (MLKs)-c-Jun amino-terminal kinase (JNK) pathways in OC formation. METHODS: We performed an OC formation assay and reverse transcription polymerase chain reaction (RT-PCR) analysis. RESULTS: We first explored the role of JNK on osteoclst formation using mouse bone marrow (BM) culture system. We found that OC formation was impaired when the JNK inhibitor was added either in early or late stage, suggesting the requirement for JNK activation during OC formation. MLKs are serine/threonine kinases that regulate signaling by the JNK. Since the JNK activity is specifically required for osteoclastogenesis, we examined the messenger ribonucleic acid (mRNA) levels of MLKs in BMs, BMMs and OCs by RT-PCR. Among MLKs, the level of MLK3 mRNA expression is highest in BMs, BMMs and OCs. Moreover, we found that the mRNA expression of MLK2 and MLK3 is increased with the differentiation of BMs to BMMs, and is sustained in OCs. Finally we investigated the role of MLK3 in OC differentiation using gene knock-down techniques. The silencing of MLK3 in BMMs partly attenuated RANKL-induced OC differentiation. CONCLUSIONS: These data suggest that JNK and MLK3 may positively regulate OC formation.
Animals ; Bone Marrow ; Gene Knockdown Techniques ; Macrophage Colony-Stimulating Factor ; Macrophages ; Mice ; Osteoclasts ; Phosphotransferases ; Polymerase Chain Reaction ; RANK Ligand ; Reverse Transcription ; RNA ; RNA, Messenger

OBJECTIVES: Osteoclasts (OCs) are bone-resorbing multinucleated cells derived from hematopoietic progenitors of the monocyte-macrophage lineage. OC precursors, such as bone marrow-derived macrophages (BMMs), are formed in the presence of macrophage colony-stimulating factor (M-CSF) and differentiate into OCs in response to M-CSF and receptor activator of nuclear factor kappaB ligand (RANKL). In this study, we investigated the role of mixed lineage kinases (MLKs)-c-Jun amino-terminal kinase (JNK) pathways in OC formation. METHODS: We performed an OC formation assay and reverse transcription polymerase chain reaction (RT-PCR) analysis. RESULTS: We first explored the role of JNK on osteoclst formation using mouse bone marrow (BM) culture system. We found that OC formation was impaired when the JNK inhibitor was added either in early or late stage, suggesting the requirement for JNK activation during OC formation. MLKs are serine/threonine kinases that regulate signaling by the JNK. Since the JNK activity is specifically required for osteoclastogenesis, we examined the messenger ribonucleic acid (mRNA) levels of MLKs in BMs, BMMs and OCs by RT-PCR. Among MLKs, the level of MLK3 mRNA expression is highest in BMs, BMMs and OCs. Moreover, we found that the mRNA expression of MLK2 and MLK3 is increased with the differentiation of BMs to BMMs, and is sustained in OCs. Finally we investigated the role of MLK3 in OC differentiation using gene knock-down techniques. The silencing of MLK3 in BMMs partly attenuated RANKL-induced OC differentiation. CONCLUSIONS: These data suggest that JNK and MLK3 may positively regulate OC formation.
Animals ; Bone Marrow ; Gene Knockdown Techniques ; Macrophage Colony-Stimulating Factor ; Macrophages ; Mice ; Osteoclasts ; Phosphotransferases ; Polymerase Chain Reaction ; RANK Ligand ; Reverse Transcription ; RNA ; RNA, Messenger

PURPOSE: Tuberculosis (TB) is a major infectious disease and is responsible for two million deaths annually. For the identification and quantitation of Mycobacterium tuberculosis (M. tuberculosis), a causative agent of TB, a sandwich enzyme-linked immunosorbent assay (ELISA) against the MPT64 protein of M. tuberculosis, an antigen marker of the M. tuberculosis complex, was developed. MATERIALS AND METHODS: The MPT64 protein was expressed, and anti-MPT64 monoclonal antibodies were prepared. A sandwich ELISA was established using recombinant MPT64 protein and anti-MPT64 monoclonal antibodies. The sandwich MPT64 ELISA was evaluated using reference and clinical mycobacterial strains. RESULTS: The sandwich MPT64 ELISA detected MPT64 protein from 2.1 ng/mL to 250 ng/mL (equivalent to 1.7x10(4) CFU/mL and 2.0x10(6) CFU/mL). All 389 clinical M. tuberculosis isolates tested positive in the sandwich MPT64 ELISA (sensitivity, 100%), and the assay showed no cross reactivity to any tested nontuberculous mycobacterial strain (specificity, 100%). CONCLUSION: The sandwich MPT64 ELISA is a highly sensitive and quantitative test for MPT64 protein, which can identify M. tuberculosis.
Antigens, Bacterial/*analysis/immunology ; Enzyme-Linked Immunosorbent Assay/*methods ; Mycobacterium tuberculosis/*immunology

Purpose: This study investigated correlations between the actual sleep time 24 hours prior to an examination and the time to achieve chloral hydrate sedation in pediatric patients. Methods: With parental consent, 84 children who were placed under moderate or deep sedation with chloral hydrate for examinations from November 19, 2020 to July 9, 2022 were recruited. Results: Patients' average age was 19.9 months. Pediatric neurology patients and those who underwent electroencephalography took significantly longer to achieve sedation with chloral hydrate. There was a negative correlation between the time to achieve sedation and actual sleep time within 24 hours prior to the examination. Positive correlations were found between the actual sleep time 24 hours prior to the examination and the second dose per weight, as well as between the sedation recovery time and awake hours before the examination. Conclusion Sleep restriction is not an effective adjuvant therapy for chloral hydrate sedation in children, and sedation effects vary according to pediatric patients' characteristics. Therefore, it would be possible to reduce the unnecessary efforts of caregivers who restrict children's sleep for examinations. It is more important to educate parents about safe sedation than about sleep restriction.

Purpose: Several predictive models have been developed to predict the pathological complete response (pCR) after neoadjuvant chemotherapy (NAC); however, few are broadly applicable owing to radiologic complexity and institution-specific clinical variables, and none have been externally validated. This study aimed to develop and externally validate a machine learning model that predicts pCR after NAC in patients with breast cancer using routinely collected clinical and demographic variables. Methods: The electronic medical records of patients with advanced breast cancer who underwent NAC before surgical resection between January 2017 and December 2020 were reviewed. Patient data from Seoul National University Bundang Hospital were divided into training and internal validation cohorts. Five machine learning techniques, including gradient boosting machine (GBM), support vector machine, random forest, decision tree, and neural network, were used to build predictive models, and the area under the receiver operating characteristic curve (AUC) was compared to select the best model. Finally, the model was validated using an independent cohort from Seoul National University Hospital. Results: A total of 1,003 patients were included in the study: 287, 71, and 645 in the training, internal validation, and external validation cohorts, respectively. Overall, 36.3% of the patients achieved pCR. Among the five machine learning models, the GBM showed the highest AUC for pCR prediction (AUC, 0.903; 95% confidence interval [CI], 0.833–0.972).External validation confirmed an AUC of 0.833 (95% CI, 0.800–0.865). Conclusion Commonly available clinical and demographic variables were used to develop a machine learning model for predicting pCR following NAC. External validation of the model demonstrated good discrimination power, indicating that routinely collected variables were sufficient to build a good prediction model.

PURPOSE: To evaluate the efficacy and safety of BOTULAX® in subjects with essential blepharospasm.METHODS: In this study, a total of 250 subjects with essential blepharospasm were enrolled at 15 investigational sites and a total of 220 subjects completed the study. The efficacy and safety were evaluated at weeks 4 and 16 after treatment compared with baseline. In total, 240 subjects were enrolled, treated with the investigational product, and evaluable for the primary efficacy assessment at week 4 after treatment; these subjects were included in the intention-to-treat (ITT) population. With the ITT set as the main efficacy set, efficacy assessment included Jankovic rating scale (JRS), functional disability score, investigator evaluation of global response and quality of life. Safety assessment including the incidence of adverse events was also performed.RESULTS: In terms of the primary efficacy endpoint (i.e., change in JRS total score at week 4 after treatment from baseline [ITT set]), mean change indicated a statistically significant reduction (p < 0.0001) and demonstrated the non-inferiority of the test drug to similar drugs. In terms of the secondary efficacy endpoints, mean change in JRS total score at week 16 after treatment and mean change in functional disability score at weeks 4 and 16 after treatment both exhibited a statistically significant reduction compared with baseline (p < 0.0001 for all). Among the 249 subjects treated with the investigational product in this study, 44 (17.67%) experienced 76 treatment emergent adverse events but no serious adverse events were observed.CONCLUSIONS: Based on the study results, BOTULAX® is considered to be an effective and safe treatment for essential blepharospasm.