Erratum: Adenosine induces intrinsic apoptosis via the PI3K/Akt/mTOR signaling pathway in human pharyngeal squamous carcinoma FaDu cells.

[This corrects the article DOI: 10.3892/ol.2018.8089.].

Adenosine induces intrinsic apoptosis via the PI3K/Akt/mTOR signaling pathway in human pharyngeal squamous carcinoma FaDu cells.

Adenosine has been identified to occur abundantly intra-and extracellularly, and to exert diverse biological functions, including the suppression of cell proliferation and the induction of apoptosis. Adenosine has been reported to induce apoptosis in several cancer cell lines; however, to the best of our knowledge, the effect of adenosine on head and neck cancer cells has not been investigated. Therefore, the purpose of the present study was to evaluate whether adenosine exerts any anticancer effect via induction of apoptosis in human pharyngeal squamous carcinoma FaDu cells. An MTT assay demonstrated that adenosine-treated FaDu cells inhibited a dose-dependent rate of cell growth, whereas human oral keratinocytes cells were unaffected by adenosine treatment. In addition, A1 and A2a adenosine receptor mRNA was detected in FaDu cells by reverse transcription-polymerase chain reaction, and adenosine-induced FaDu cell death was significantly suppressed by treatment with ATL-444, an antagonist of these receptors. Furthermore, adenosine-induced cell growth inhibition was exerted via apoptosis, as confirmed by the analysis of DNA fragmentation, Hoechst nuclear staining and flow cytometry with Annexin V-fluorescein isothiocyanate and propidium iodide staining. Adenosine was also demonstrated to induce an increase in Bcl-associated X expression, a decrease in B-cell lymphoma 2 expression, the release of cytochrome c from mitochondria, and the activation of caspase-3, -9 and poly(ADP-ribose) polymerase in FaDu cells. Finally, phosphoinositide 3-kinase (PI3K), RAC serine/threonine-protein kinase (Akt) and mechanistic target of rapamycin (mTOR) phosphorylation was found to be significantly inhibited in adenosine-treated FaDu cells, as was phosphorylation of the mTOR downregulators, S6 kinase ß1, eukaryotic translation initiation factor 4E-binding protein 1, and eukaryotic translation initiation factor 4 γ1. Taken together, these results indicate that adenosine induces apoptosis via the mitochondrial intrinsic pathway, and activates caspase-3 and -9 activity via the PI3K/Akt/mTOR signaling pathway.

In Vivo and In Vitro Anti-Inflammatory Effects of Aqueous Extract of Anthriscus sylvestris Leaves.

Anthriscus sylvestris (L.) Hoffm. is a common perennial herb that is widely distributed in Europe, Korea, and New Zealand. The root of A. sylvestris has been used in Korean traditional medicine as an antitussive and cough remedy. However, the physiologically active function of A. sylvestris leaves is not yet known. In this study, we evaluated the anti-inflammatory effects, as well as the underlying molecular mechanisms of an aqueous extract of A. sylvestris leaves (AE-ASL) in vitro and in vivo. Our results indicated that pretreatment with AE-ASL significantly inhibited the lipopolysaccharide (LPS)-induced secretion of nitric oxide (NO) and prostaglandin E2 in RAW264.7 cells, without showing cytotoxicity. In addition, the LPS-induced mRNA and protein expression of inducible NO synthase, cyclooxygenase-2, and inflammatory mediators such as tumor necrosis factor alpha interleukin (IL)-1ß, and IL-6 was attenuated by pretreatment with AE-ASL in a dose-dependent manner. Therefore, we investigated the activation of nuclear factor (NF)-κB, a transcription factor regulating the expression of inflammation-related genes. AE-ASL inhibited the nuclear translocation of the NF-κB p65 subunit by suppressing the phosphorylation and degradation of the inhibitor of NF-κB (IκBα). Further, AE-ASL inhibited the LPS-induced phosphorylation of mitogen-activated protein kinases (MAPKs) in RAW264.7 cells. Orally administered AE-ASL (50, 100, and 200 mg/kg of body weight [BW]) suppressed the development of carrageenan-induced rat paw edema by 15%, 31%, and 40%, respectively, after 4 h. Altogether, our results suggest that AE-ASL possesses anti-inflammatory activity, based on the suppression of NF-κB and MAPK pathways in vitro and inhibition of the carrageenan-induced paw edema in vivo.

Aqueous extract of Codium fragile alleviates osteoarthritis through the MAPK/NF-κB pathways in IL-1ß-induced rat primary chondrocytes and a rat osteoarthritis model.

BACKGROUND: Codium fragile (Suringar) Hariot has been used as a potential remedy in traditional medicine because of its anti-inflammatory and anti-oxidant effects. Osteoarthritis is a chronic progressive joint disease, characterized by complex mechanisms related to inflammation and degeneration of articular cartilage. In this study, we aimed to evaluate the cartilage protective effect of an aqueous extract of Codium fragile (AECF) using rat primary chondrocytes and the osteoarthritis animal model induced by destabilization of the medial meniscus (DMM). METHODS: In vitro, rat primary cultured chondrocytes were pre-treated with AECF (0.5, 1, and 2mg/mL) for 1h and then incubated with interleukin-1ß (10ng/mL) for 24h. Nitrite production was detected by the Griess reagent. Alteration of the protein levels of iNOS, MMP-13, ADAMTS-4, ADAMTS-5, mitogen-activated protein kinases (MAPKs), and nuclear factor-κB (NF-κB) was detected by western blotting. In vivo, osteoarthritis was induced by DMM of Sprague Dawley (SD) rats. The rats subjected to destabilization of the medial meniscus (DMM) surgery were orally administered with AECF (50, 100, and 200mg/kg bodyweight) or distilled water for 8w. The severity of cartilage lesions was evaluated by safranin O staining and the Osteoarthritis Research Society International (OARSI) score. RESULTS: These results demonstrated that AECF significantly inhibited nitrite production and inhibited the levels of iNOS, MMP-13, ADAMTS-4, and ADAMTS-5 in interleukin-1ß-induced rat primary cultured chondrocytes. Moreover, AECF suppressed interleukin-1ß-induced NF-κB activation in the nucleus and phosphorylation of ERK1/2 and JNK in the cytosol. In vivo, the cartilage lesions in AECF-treated osteoarthritis rats exhibited less proteoglycan loss and lower OARSI scores. CONCLUSIONS: These results suggested that AECF is a potential therapeutic agent for the alleviation of osteoarthritis progression.

Aqueous extract of Codium fragile suppressed inflammatory responses in lipopolysaccharide-stimulated RAW264.7 cells and carrageenan-induced rats.

Codium fragile (Suringar) Hariot has been used in Oriental medicine for the treatment of enterobiasis, dropsy, and dysuria and has been shown to have various biological effects. In this study, we evaluated the anti-inflammatory effects of aqueous extract of C. fragile (AECF) using in vitro and in vivo models. Nitric oxide (NO), prostaglandin E2 (PGE2), inflammatory-related mRNAs, and proteins were determined using the Griess assay, enzyme-linked immunosorbent assay (ELISA), reverse transcription-polymerase chain reaction (RT-PCR), and western blotting, respectively. Our results indicate that pretreatment of cells with AECF (50, 100 and 200µg/mL) significantly inhibited LPS-induced secretion of NO and PGE2 in RAW264.7 cells without cytotoxicity. We also found that AECF (100 and 200µg/mL) inhibited LPS-induced inducible NO synthase (iNOS) and cyclooxygenase (COX)-2 expression in a dose-dependent manner. Additionally, pretreatment of cells with AECF (100 and 200µg/mL) inhibited LPS-induced production of inflammatory cytokines including tumor necrosis factor (TNF)-α, interleukin (IL)-1ß, and IL-6. It also prevented the nuclear translocation of nuclear factor (NF)-κB by suppressing the phosphorylation and degradation of inhibitor of NF-κB (IκB)-α. Furthermore, AECF (100 and 200µg/mL) inhibited the phosphorylation of the mitogen-activated protein kinases (MAPKs) extracellular signal-regulated kinase (ERK) 1/2, c-Jun N-terminal kinase (JNK), and p38. In addition, orally administered 50, 100, and 200mg/kg body weight of AECF dose-dependently suppressed carrageenan-induced rat paw edema thickness by 6%, 31%, and 50% respectively, after 4h. Furthermore, the anti-inflammatory effect was comparable to that observed in animals treated with the standard drug diclofenac sodium (56%) in vivo. Collectively, our results suggest that AECF exerts potential anti-inflammatory effects by suppressing NF-κB activation and MAPKs pathways in vitro, as well as inhibiting carrageenan-induced rat paw edema thickness in vivo. These findings indicate that AECF could be further developed as an anti-inflammatory drug.

Outbreak of Ciprofloxacin-Resistant Shigella sonnei Associated with Travel to Vietnam, Republic of Korea.

We investigated an October 2014 outbreak of illness caused by Shigella sonnei in a daycare center in the Republic of Korea (South Korea). The outbreak strain was resistant to extended-spectrum cephalosporins and fluoroquinolones and was traced to a child who had traveled to Vietnam. Improved hygiene and infection control practices are needed for prevention of shigellosis.

Clinical characteristics of lung abscess in children: 15-year experience at two university hospitals.

PURPOSE: Information on the clinical features of lung abscess, which is uncommon in children, at hospitalizationis helpful to anticipate the disease course and management. There is no report concerning lung abscess in Korean children. We aimed to identify the clinical characteristics of pediatric lung abscess and compare the difference between primary and secondary abscess groups. METHODS: The medical records of 11 lung abscess patients (7 males and 4 females) from March 1998 to August 2011 at two university hospitals were retrospectively reviewed. The clinical characteristics, symptoms, underlying disease, laboratory and radiologic findings, microbiological results, and treatments were examined. RESULTS: Six patients had underlying structural-related problems (e.g., skeletal anomalies). No immunologic or hematologic problem was recorded. The mean ages of the primary and secondary groups were 2.4 and 5.3 years, respectively, but the difference was not statistically significant. The mean length of hospital stay was similar in both groups (22.8 days vs. 21.4 days). Immunologic studies were performed in 3 patients; the results were within the normal range. Most patients had prominent leukocytosis. Seven and 4 patients had right and left lung abscess, respectively. Staphylococcus aureus, Streptococcus pneumoniae, and antimycoplasma antibodies were detected in both groups. Two patients with primary lung abscess were administered antibiotics in the absence of other procedures, while 8 underwent interventional procedures, including 5 with secondary abscess. CONCLUSION: The most common symptoms were fever and cough. All patients in the primary group were younger than 3 years. Structural problems were dominant. Most patients required interventional procedures and antibiotics.

Purification and characterization of a novel fibrinolytic α chymotrypsin like serine metalloprotease from the edible mushroom, Lyophyllum shimeji.

A novel fibrinolytic enzyme was purified from Lyophyllum shimeji, a popular edible mushroom in Asia. The enzyme was purified using combination of anion exchange chromatography on a Mono Q 5/5 column and size exclusion gel filtration chromatography on Superdex 200 100/300 column. This purification protocol resulted 80.9-fold purification of the enzyme and a final yield of 5.7%. The molecular weight of the purified enzyme was estimated to be 21 kDa by SDS-PAGE and size exclusion gel filtration. The N-terminal amino acid sequence was found to be ITFQSASP, which is dissimilar from that of known fibrinolytic enzymes. The purified enzyme was a neutral protease with an optimal reaction pH and temperature of 8.0 and 37°C, respectively. Enzymatic activity was inhibited by Cu(2+) and Co(2+). It was also significantly inhibited by PMSF and TPCK. Furthermore, it was found to exhibit a higher specificity for S-7388, a well-known chymotrypsin chromogenic substrate, indicating chymotrypsin like serine metalloprotease. The relative fibrinolytic activity of 5 µg purified enzyme have two fold more activity than 1 unit/ml of plasmin on fibrin plate. Furthermore, purified enzyme preferentially hydrolyzed the Aα-chain followed by the Bß- and γ-chain of fibrinogen, which is precursor of fibrin. Therefore, these data suggests that the fibrinolytic enzyme derived from edible mushroom, L. shimeji, might be useful for thrombolytic therapy and preventing thrombotic disease.

Anticancer activity of Saussurea lappa extract by apoptotic pathway in KB human oral cancer cells.

CONTEXT: Saussurea lappa Dence (Compositae) is used as a traditional herbal medicine to treat abdominal pain and tenesmus in East Asia. Current studies have shown that S. lappa has anticancer activity in divergent of cancer cells. However, the effects of S. lappa on oral cancer and its mechanisms of action have yet to be elucidated. OBJECTIVE: To explore its potential chemotherapeutic effects and mechanism of cell growth inhibition on human oral cancer cells. MATERIALS AND METHODS: The dried roots of S. lappa were used in this study. Cell viability of KB cells was evaluated by 3-[4, 5-dimethylthiazol-2-yl]-2, 5-diphenyltetrazolium bromide assay after treatment with 30 µg/ml of methanol extract from the dried roots of S. lappa. To understand whether its effect on cell death is related with apoptosis pathway, we performed DNA fragmentation assay, western blot, caspase activity assay and fluorescence-activated cell sorting (FACS) analysis. RESULTS: Treatment of S. lappa extract onto KB cells reduced cell viability significantly with an IC50 value of 30 µg/ml. The formation of a DNA ladder was observed starting at the 24 h treatment. In western blotting analysis, the S. lappa extract induced the proteolytic processing of caspase-3, -9 and poly (ADP-ribose) polymerase, a significant increase of Bax and marked reduction of Bcl-2. We also confirmed the activation of caspase-3/-7 in living KB cells by fluorescence microscopy. CONCLUSION: These results suggested that S. lappa extract inhibited cell proliferation through the apoptosis pathway in KB human oral cancer cells.

Clinical characteristics of labyrinthine concussion.

BACKGROUND AND OBJECTIVES: Inner ear symptoms like hearing loss, dizziness or tinnitus are often developed after head trauma, even in cases without inner ear destruction. This is also known as labyrinthine concussion. The purpose of this study is to determine the clinical manifestations, characteristics of audiometry and prognostic factors of these patients. MATERIALS AND METHODS: We reviewed the medical records of the 40 patients that had been diagnosed as labyrinthine concussion from 1996 to 2007. We studied the hearing levels in each frequency and classified them according to type and degree of hearing loss. Rates of hearing improvement were evaluated according to age, sex, hearing loss type, degree and presence of dizziness or tinnitus. To find out any correlation between hearing improvement and these factors, we used χ(2) test or Fisher's exact test. RESULTS: Bilateral hearing loss was observed in 22 patients, and unilateral hearing loss in 18 patients. There were 4 (6.5%) ascending, 34 (54.8%) descending, 24 (38.7%) flat type hearing loss, which indicated hearing loss was greater in high frequencies than low frequencies. Among 62 affected ears, 20 (32.3%) gained improvement, and it was achieved mainly in low frequencies. There were only 2 ears with dizziness in 20 improved ears and among 20 dizziness accompanied ears, also only 2 ears were improved. CONCLUSIONS: High frequencies are more vulnerable to trauma than low frequencies. The hearing gain is obtained mainly in low frequencies, and association with dizziness serves poor prognosis.

Isolation of 151 mutants that have developmental defects from T-DNA tagging.

In order to understand the mechanisms underlying plant development, a necessary first step involves the elucidation of the functions of the genes, via the analysis of mutants that exhibit developmental defects. In this study, an activation tagging mutant library harboring 80,650 independent Arabidopsis transformants was generated in order to screen for developmental mutants. A total of 129 mutants manifesting dominant developmental abnormalities were isolated, and their T-DNA insertion loci were mapped. The activation of one or more genes adjacent to a T-DNA insertion locus was confirmed in eight dominant mutants. A gene adjacent to the right border was usually activated by the 35S enhancers. Interestingly, the transcriptional activation of multiple genes within a broad range was observed in one of the mutants, which raises the possibility that activation by the 35S enhancers was not limited strictly to a single gene. In order to gain a better understanding of sexual reproduction in higher plants, we isolated 22 mutants exhibiting defects in female gametophyte development, and determined their T-DNA insertion loci. We propose that this mutant population may prove useful in the further determination of the functions of genes that play important roles in plant development.

The 35S promoter used in a selectable marker gene of a plant transformation vector affects the expression of the transgene.

Positive selection of transgenic plants is essential during plant transformation. Thus, strong promoters are often used in selectable marker genes to ensure successful selection. Many plant transformation vectors, including pPZP family vectors, use the 35S promoter as a regulatory sequence for their selectable marker genes. We found that the 35S promoter used in a selectable marker gene affected the expression pattern of a transgene, possibly leading to a misinterpretation of the result obtained from transgenic plants. It is likely that the 35S enhancer sequence in the 35S promoter is responsible for the interference, as in the activation tagging screen. This affected expression mostly disappeared in transgenic plants generated using vectors without the 35S sequences within their T-DNA region. Therefore, we suggest that caution should be used in selecting a plant transformation vector and in the interpretation of the results obtained from transgenic approaches using vectors carrying the 35S promoter sequences within their T-DNA regions.