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Mechanism Underlying Shikonin-induced Apoptosis and Cell Cycle Arrest on SCC25 Human Tongue Squamous Cell Carcinoma Cell Line.

Sang Hun OH ; Sung Jin PARK ; Su Bin YU ; Yong Ho KIM ; In Ryoung KIM ; Bong Soo PARK

International Journal of Oral Biology.2015;40(1):51-61. doi:10.11620/IJOB.2015.40.1.051

Shikonin, a major ingredient in the traditional Chinese herb Lithospermumerythrorhizon, exhibits multiple biological functions including antimicrobial, anti-inflammatory, and antitumor effects. It has recently been reported that shikonin displays antitumor properties in many cancers. This study was aimed to investigate whether shikonin could inhibit oral squamous carcinoma cell (OSCC) growth via mechanisms of apoptosis and cell cycle arrest. The effects of shikonin on the viability and growth of OSCC cell line, SCC25 cells were assessed by MTT assay and clonogenic assays, respectively. Hoechst staining and DNA electrophoresis indicated that the shikonin-treated SCC25 cells were undergoing apoptosis. Western blotting, immunocytochemistry, confocal microscopy, flow cytometry, MMP activity, and proteasome activity also supported the finding that shikonin induces apoptosis. Shikonin treatment of SCC25 cells resulted in a time- and dose-dependent decrease in cell viability, inhibition of cell growth, and increase in apoptotic cell death. The treated SCC25 cells showed several lines of apoptotic manifestation as follows: nuclear condensation; DNA fragmentation; reduced MMP and proteasome activity; decrease in DNA contents; release of cytochrome c into cytosol; translocation of AIF and DFF40 (CAD) onto the nuclei; a significant shift in Bax/Bcl-2 ratio; and activation of caspase-9, -7, -6, and -3, as well as PARP, lamin A/C, and DFF45 (ICAD). Shikonin treatment also resulted in down-regulation of the G1 cell cycle-related proteins and up-regulation of p27(KIP1). Taken together, our present findings demonstrate that shikonin strongly inhibits cell proliferation by modulating the expression of the G1 cell cycle-related proteins, and that it induces apoptosis via the proteasome, mitochondria, and caspase cascades in SCC25 cells.
Apoptosis* ; Asian Continental Ancestry Group ; Blotting, Western ; Carcinoma, Squamous Cell* ; Caspase 9 ; Cell Cycle Checkpoints* ; Cell Death ; Cell Line* ; Cell Proliferation ; Cell Survival ; Cytochromes c ; Cytosol ; DNA ; DNA Fragmentation ; Down-Regulation ; Electrophoresis ; Flow Cytometry ; Humans ; Immunohistochemistry ; Microscopy, Confocal ; Mitochondria ; Proteasome Endopeptidase Complex ; Tongue* ; Up-Regulation

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Identification of the Bacteria Isolated from Oral Cavities in Korea.

Mi Hwa CHOI ; Soon Nang PARK ; Joong Ki KOOK

International Journal of Oral Biology.2015;40(1):41-50. doi:10.11620/IJOB.2015.40.1.041

The aim of this study was to identify bacteria isolated from the oral cavities and to determine their antimicrobial susceptibility against eight antibiotics. The bacterial strains were obtained from the Korean Collection for Oral Microbiology (KCOM). The bacteria were identified by comparing 16S rDNA sequences at the species level. The data showed that 77 bacterial strains were predominantly identified as streptococci (49.4%) and staphylococci (14.3%). Minimum inhibitory concentrations (MIC) were determined using a broth dilution assay to test the sensitivity of the bacterial strains. The MIC values of the oral bacterial strains against antibiotics were different. Streptococci were sensitive to clindamycin, cefuroxime axetil, and vancomycin, and they were resistant to tetracycline. Staphylococci also were sensitive to clindamycin, cefuroxime axetil, and vancomycin, and they were resistant to penicillin antibiotics. Gram-negative bacterial strains were sensitive to tetracycline and were resistant to clindamycin. These results suggest that the antimicrobial susceptibility test is necessary in deciding the prescription for antibiotics, to prevent the misuse or abuse of antibiotics.
Anti-Bacterial Agents ; Bacteria* ; Cefuroxime ; Clindamycin ; DNA, Ribosomal ; Korea ; Microbial Sensitivity Tests ; Penicillins ; Prescriptions ; Tetracycline ; Vancomycin

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Effects of Sub Minimal Inhibitory Concentration of Metronidazole and Penicillin on Morphology of Aggregatibacter actinomycetemcomitans: Scanning Electron Microscopy Observation.

Ye Won KWON ; Si Young LEE

International Journal of Oral Biology.2015;40(1):35-39. doi:10.11620/IJOB.2015.40.1.035

Minimal inhibitory concentration (MIC) is the lowest concentration of antibiotics that inhibits the visible growth of bacteria. It has been reported that sub-MIC of antibiotics may result in morphological alterations, along with the biochemical and physiological changes in bacteria. The purpose of this study was to examine morphological changes of Aggregatibacter actinomycetemcomitans, after the treatment with sub-MIC metronidazole and penicillin. The bacterial morphology was observed with scanning electron microscope, after incubating with sub-MIC antibiotics. The length of A. actinomycetemcomitans was increased after the incubation with sub-MIC metronidazole and penicillin. Sub-MIC metronidazole and penicillin inhibited bacterial division and induced long filaments. Our study showed that metronidazole and penicillin can induce the morphological changes in A. actinomycetemcomitans.
Aggregatibacter actinomycetemcomitans* ; Anti-Bacterial Agents ; Bacteria ; Metronidazole* ; Microscopy, Electron, Scanning* ; Penicillins*

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The Effect of Autogenous Demineralized Dentin Matrix and Interleukin-6 on bone Regeneration.

Won Seok JANG ; Min Gu KIM ; Dae Suk HWANG ; Gyoo Cheon KIM ; Uk Kyu KIM

International Journal of Oral Biology.2017;42(4):203-211. doi:10.11620/IJOB.2017.42.4.203

The aim of this study was to evaluate the role of demineralized and particulate autogenous tooth, and interleukin-6 in bone regeneration. A demineralized and particulate autogenous tooth was prepared and human osteoblast-like cells (MG63) and human osteosarcoma cells were inoculated into the culture. The rate of cell adhesion, proliferation and mineralization were examined, and the appearance of cellular attachment was observed. An 8 mm critical size defect was created in the cranium of rabbits. Nine rabbits were divided into three groups including: An experimental group A (3 rabbits), in which a demineralised and particulate autogenous tooth was grafted; an experimental group B (3 rabbits), in which a demineralized, particulate autogenous tooth was grafted in addition to interleukin-6 (20 ng/mL); and a control group. The rabbits were sacrificed at 1, 2, 4 and 6 weeks for histopathological examination with H-E and Masson's Trichrome, and immunohistochemistry with osteocalcin. The cell-based assay showed a higher rate of cell adhesion, mineralization and cellular attachment in the experimental group A compared with the control group. The animal study revealed an increased number of osteoclasts, newly formed and mature bones in the experimental group A compared with the control group. Eventually, a higher number of osteoclasts were observed in the experimental group B. However, the emergence of newly formed and mature bone was lower than in the experimental group A. The current results suggest that treatment with demineralized and particulate autogenous tooth and interleukin-6 is not effective in stimulating bone regeneration during the bone grafting procedure.
Animals ; Bone Regeneration* ; Bone Transplantation ; Cell Adhesion ; Dentin* ; Humans ; Immunohistochemistry ; Interleukin-6* ; Miners ; Osteocalcin ; Osteoclasts ; Osteosarcoma ; Rabbits ; Skull ; Tooth ; Transplants

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Capillary Electrophoresis Detection of Hydrogen Peroxide by Using Titanium Ion and 4-(2-thiazolylazo)resorcinol.

Dong Vu PHUONG ; Hoon YOO

International Journal of Oral Biology.2017;42(4):197-201. doi:10.11620/IJOB.2017.42.4.197

A novel method for the detection of hydrogen peroxide in aqueous solution was developed via reaction between H₂O₂, trivalent titanium ion (Ti³⁺) and 4-(2-thiazolylazo) resorcinol (TAR), resulting in a ternary complex with a maximum UV absorbance at 530 nm. The CE detection of H₂O₂ was fast, sensitive and cost-effective without pretreatment procedures. H₂O₂ was detected within 15 min at 1 to 100 µM range with the lowest detection limit at 1.0 µM. Under the optimized CE conditions, the concentration of H₂O₂ in coffee or tea extract was quantitatively determined. Our results show that CE detection of the ternary complex of H₂O₂-Ti³⁺-TAR has potential applications for the detection of H2O2 in aqueous sources.
Capillaries* ; Coffee ; Electrophoresis, Capillary* ; Hydrogen Peroxide* ; Hydrogen* ; Limit of Detection ; Methods ; Tea ; Titanium*

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Regulation of Inflammatory Response in Periodontal Ligament Cells by Transglutaminase 2.

Sun Young LEE ; Cheol Hun JANG ; Je Hwang RYU

International Journal of Oral Biology.2017;42(4):191-196. doi:10.11620/IJOB.2017.42.4.191

Transglutaminase2 (TGM2) is a multi-functional calcium dependent enzyme that affects angiogenesis, apoptosis, differentiation, attachment, and changes in the extracellular matrix. However, its function in periodontal tissue has not yet been studied. The aim of this study was to investigate the association of the TGM2 expression and the modulation of inflammatory mediators in inflamed periodontal ligament (PDL) cells induced by pro-inflammatory cytokines such as Interleukin-1β and the Tumor necrosis factor-α. The expression of TGM2 was increased in the inflamed periodontal tissue and PDL cells. Over-expressed TGM2 in the PDL cells increased expression of MMP1, MMP3, IL-6, CXCL8, and PTGS2. Conversely, inhibition of TGM2 activity using LDN27219, a TGM2 inhibitor, resulted in decreased expression of MMP1, MMP3, IL-6, and CXCL8. The mRNA expression was confirmed by RT-PCR and quantified by qRT-PCR. Protein levels were also confirmed by immunofluoroscence staining. These results suggest that TGM2 plays an important role in the regulation of inflammatory mediators which exacerbate tissue damage in inflamed periodontal tissue.
Apoptosis ; Calcium ; Cyclooxygenase 2 ; Cytokines ; Extracellular Matrix ; Inflammation ; Interleukin-6 ; Necrosis ; Periodontal Ligament* ; Periodontitis ; RNA, Messenger

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Latex of Ficus carica L. Induces Apoptosis Through Caspase and Bcl-2 Family in FaDu Human Hypopharynx Squamous Carcinoma Cells.

Bo Su SHIN ; Seul Ah LEE ; Sung Min MOON ; Seul Hee HAN ; Eun Ju HWANG ; Su Gwan KIM ; Do Kyung KIM ; Jin Soo KIM ; Bo Ram PARK ; Chun Sung KIM

International Journal of Oral Biology.2017;42(4):183-190. doi:10.11620/IJOB.2017.42.4.183

Ficus carica L. (common fig), one of the first plants cultivated by humans, originated in the Mediterranean basin and currently grows worldwide, including southwest Asia and South Korea. It has been used as a traditional medicine for treatment of metabolic, cardiovascular, and respiratory diseases as well as hemorrhoids and skin infections. Its pharmacological properties have recently been studied in detail, but research on the anti-cancer effect of its latex has been only been studied on a limited basis on several cell lines, such prostate cancer, breast cancer, and leukemia. In this study, we investigated the anti-cancer activity of the latex of Ficus carica L.and its underlying mechanism in FaDu human hypopharynx squamous carcinoma cells. (See Ed. note above) We confirmed through SDS-PAGE analysis and gelatinolytic activity analysis that the latex of Ficus carica contains cysteine protease ficin. Our data showed that the latex inhibited cell growth in a dose-dependent manner. In addition, the latex treatment markedly induced apoptosis in FaDu cells as determined by FACS analysis, elevated expression level of cleaved caspase-9, -3 and PARP (poly (ADP-ribose) polymerase), and. increased the expression of Bax (pro-apoptotic factor) while decreasing the expression of Bcl-2 (anti-apoptotic factor). Taken together, these results suggested that latex containing the ficin inhibited cell growth and induced apoptosis by caspase and the Bcl-2 family signaling pathway in FaDu human hypopharynx squamous carcinoma cells. These findings point to the potential of latex of Ficus carica to provide a novel chemotherapeutic drug due to its growth inhibition effects and induction of apoptosis in human oral cancer cells.
Apoptosis* ; Asia ; Breast Neoplasms ; Carcinoma, Squamous Cell* ; Carica* ; Caspase 9 ; Cell Line ; Cysteine Proteases ; Electrophoresis, Polyacrylamide Gel ; Ficain ; Ficus* ; Hemorrhoids ; Humans ; Humans* ; Hypopharynx* ; Korea ; Latex* ; Leukemia ; Medicine, Traditional ; Mouth Neoplasms ; Prostatic Neoplasms ; Skin

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The Effect of 1-Propanol on the Rotational Mobility of n-(9-Anthroyloxy) stearic acid in Outer Monolayers of Neuronal and Model Membranes.

Tae Young AHN ; Seong Deok JIN ; Hak Jin YANG ; Chang Dae YOON ; Mi Kyung KIM ; Taek Kyung AN ; Young Jun BAE ; Sang Jin SEO ; Gwon Su KIM ; Moon Kyoung BAE ; Soo Kyoung BAE ; Hye Ock JANG

International Journal of Oral Biology.2017;42(4):175-181. doi:10.11620/IJOB.2017.42.4.175

The aim of this study was to provide a basis for the molecular mechanism underlying the pharmacological action of ethanol. We studied the effects of 1-propanol on the location of n-(9-anthroyloxy)palmitic acid or stearic acid (n-AS) within the phospholipids of synaptosomal plasma membrane vesicles (SPMV). The SPMV were isolated from the bovine cerebral cortex and liposomes of total lipids (SPMVTL) and phospholipids (SPMVPL). 1-Propanol increased the rotational mobility of inner hydrocarbons, while decreasing the mobility of membrane interface, in native and model membranes. The degree of rotational mobility varied with the number of carbon atoms at positions 16, 12, 9, 6 and 2 in the aliphatic chain of phospholipids in the neuronal and model membranes. The sensitivity of increasing or decreasing rotational mobility of hydrocarbon interior or surface by 1-propanol varied with the neuronal and model membranes in the following order: SPMV, SPMVPL and SPMVTL.
1-Propanol* ; Carbon ; Cell Membrane ; Cerebral Cortex ; Ethanol ; Hydrocarbons ; Liposomes ; Membranes* ; Neurons* ; Phospholipids

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Distribution of Cold Receptor Transient Receptor Potential Melastatin 8-Immunopositive Axons in the Mouse Dental Pulp and Periodontal Tissue.

Tae Heon KIM ; Jae Sik LEE ; Yun Sook KIM ; Yong Chul BAE

International Journal of Oral Biology.2017;42(4):169-174. doi:10.11620/IJOB.2017.42.4.169

Transient receptor potential melastatin 8 (TRPM8) plays a crucial role in innocuous cool sensation, acute cold pain and cold-induced hyperalgesia during pathologic conditions. To help understand TRPM8-mediated cold perception in the dental pulp and periodontal tissues, we examined the distribution of TRPM8-immunopositive (+) axons in molar and incisor pulp and periodontal tissues using transgenic mice expressing a genetically encoded axonal tracer in TRPM8+ neurons. In the radicular pulp of the molar teeth, a small number of TRPM8+ axons were observed. TRPM8+ axons branched frequently and extensively in the core of coronal pulp, forming a network in the peripheral pulp. Some TRPM8+ axons ascended between odontoblasts and were observed in the dentinal tubule. TRPM8+ axons were linear-shaped in the radicular pulp, whereas many TRPM8+ axons showed portions shaped like beads connected with thin axonal stands at the peripheral pulp. TRPM8 was densely expressed in the bead portions. In the incisor pulp, TRPM8+ axons were occasionally observed in the core of the coronal pulp and rarely observed at the peripheral pulp. TRPM8+ axons were occasionally observed and showed a linear shape rather than a bead-like appearance in the periodontal ligament and lamina propria of the gingival tissue. These findings, showing differential distribution of TRPM8+ axons between radicular and coronal portions of the molar pulp, between incisor and molar pulp, and between dental pulp and periodontal tissues, may reflect differential cold sensitivity in these regions.
Animals ; Axons* ; Dental Pulp* ; Dentin ; Hyperalgesia ; Immunohistochemistry ; Incisor ; Mice* ; Mice, Transgenic ; Molar ; Mucous Membrane ; Neurons ; Odontoblasts ; Periodontal Ligament ; Sensation ; Tooth

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Icaritin, a Flavonoid Derived from the Herb Epimedium, Promotes Osteogenic Differentiation of MC3T3-E1 Cells.

Dan Bi PARK ; Hee Su LEE ; Seong Hee KO

International Journal of Oral Biology.2017;42(4):163-168. doi:10.11620/IJOB.2017.42.163

Osteoporosis is a metabolic bone disease that is characterized by low bone mass resulting from an increase in bone resorption relative to bone formation. The most current therapies for osteoporosis have focused on inhibiting bone resorption by osteoclasts. The purpose of this study is to develop new anabolic agents for treatment of osteoporosis that have fewer risks compared to conventional therapies. We searched the natural products that were derived from the traditional Asian medicines which have been used for treatment of bone related diseases. Icaritin is a flavonoid glycoside derived from the herb Epimedium which has beneficial effects on bone formation. To determine the effect of icaritin on bone formation, we examined the effect of icaritin on MC3T3-E1 cell proliferation and differentiation. For determining the effects of icaritin on proliferation, we performed the MTT assay using MC3T3-E1 cells. To evaluate whether icaritin could promote the osteogenic differentiation of MC3T3-E1 cells, alkaline phosphatase (ALP) activity and mRNA expressions of Runx2, osteocalcin (OCN), RANKL, and osteoprotegerin (OPG) were determined. Icaritin increased MC3T3-E1 cell proliferation. Icaritin increased the ALP activity of MC3T3-E1 cells on 72 hour culture in osteogenic media. mRNA expression of Runx2 was increased after 24 hour culture with icaritin. mRNA expression of osteocalcin was increased after 72 hour culture with icaritin. In addition, icaritin increased the mRNA expressions of OPG and RANKL. However, icaritin increased the mRNA expression of OPG much more than that of RANKL, and then, it increased the OPG/RANKL ratio. These results suggest that icaritin promotes osteogenic differentiation of osteoblasts and decreases osteoclast formation regulated by osteoblasts.
Alkaline Phosphatase ; Anabolic Agents ; Asian Continental Ancestry Group ; Biological Products ; Bone Diseases, Metabolic ; Bone Resorption ; Cell Proliferation ; Epimedium* ; Humans ; Osteoblasts ; Osteocalcin ; Osteoclasts ; Osteogenesis ; Osteoporosis ; Osteoprotegerin ; RNA, Messenger

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